NATIONAL INSTITUTE OF GENETICSJAPANANNUAL REPORT
No. 8

1957

Published by

THE NATIONAL INSTITUTE OF GENETICS
Misima. Sizuoka-ken. Japan1958

Annual Reportof theNational Institute of Genetics
No.8,

1957

Published by

The National Institute of Genetics, Japan
1958

t

CONTENTS,

General statement
Abstracts of diary of 1957
Staff
,
,
,
, .. .. .. .. .. .. .. ..
CouncilResearch program for 1957.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Foreign visitors in 1957
Researches carried out in 1957 .. . . . . . . . . . . . . .. .. . . . . . . . . . . . . . . . . . . . .

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2
3
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8
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A. Genetics, biochemistry and cytology of insects . . . . . . . . . . . . . . . .
1. Inheritance of a new mutant "brown-3" of the silkworm(TAZIMA) .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. ..
2. Studies on a food preference mutant in the silkworm, VIII.
Selection for higher incidence of non-preference individuals
and their genetic analysis (TAZIMA and MACHIDA)
3. The resistance of strains of D. melanogaster to DDT and
Dieldrin (OSHIMA)...... .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. ....
4. Genetical and biochemical studies on pterine oxidation in Drosophila melanogaster (NAWA, TAIRA and SAKAGUCHI) ........
5. Purine Catabolism in D. melanogaster (MORITA and OSHIMA) ..
6. Dopa oxidase in the eye-color mutants of Drosophila melanogaster (TAIRA and NAWA)
7. Formation of Drosophila eye pigments (TAIRA and NAWA) .. ..
8. Genetic determination of tyrosinase and protyrosinase in bloodof silkworm, Bombyx mori L. (SAKAGUCHI)
9. Chromosomal constitution of the original Nl 2 strain of thesilkworm (TSUJITA) .... .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. ....
10. Studies on the mitochondria in the gland cells of Malpighiantubules of the silkworm larvae by means of ultra-thin sections (TSUJITA) .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. ....

9

B. Cytogenetics Genetics, Cytology and Biochemistry of tumors ..
11. Two chemicals promoting cell divisions in the Yoshida sarcoma(OGAWA, FUJIOKA and ABE)
12. Biochemical similarity of mitotic stimulants in normal andsarcoma cells of rat (OGAWA) .. .. .. .. .. .. .. .. .. .. .. .. .. ....
13. Idiogram of a new subline of the Yoshida sarcoma and someof its properties (Yosida) .. . . .. . . .. .. .. .. .. .. .. .. .. .. .. ....

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10
11
12
13
14
14
16
18

19
20
20
22
24

CONTENTS

ii

14.

A karyological study on the Yoshida sarcoma cells infiltrated
into organs of a tumor-bearing rat (Yoshida) .. .. .. .. .. .. .. 24
15. Karyological study on the normal somatic and malignant
tumor cells in the human (Yoshida and Tabata)25
16. Change in the transplantability of the MY-mouse sarcoma in
relation to the change of the chromosome number (Yosida
and Hamada) ... . . . . . . . . . . . . . .. .. . . . . . . . . . . . . . . . . . . .. . . . . 27
17. Effect of 8-azaguanine and its related compounds on the chromosomes of the Yoshida sarcoma cells (Yosida and Hirumi)
27
18. Effect of chemical anions on the spreading and splitting ofanimal chromosomes (Yosida and Tabata) ..... . . . . . . . .. .. 28
C. Developmental genetics of Animals.. . . . . . . . . . . . . . . . . . . . . . . . . . . 29
19. Development of muscle protein in early chick embryo (OGAWA,
KAWAHARA and MIURA)
29
20. Promoting effect of gonade extracts on the spawning of thepearl oyster (Pinctada Martensii) (OGAWA, HARADA and AI).. 31
D. Cytology of Animals.. . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . .
21. Mitotic activity of regenerating liver cells after partial hepatectomy (OGAWA)

32

E. Genetics and cytology of cereal crops . . . . . . . . . . . . .. . . . . . . .. . .
22. Ecogenetical studies in Agropyron (SAKAMOTO).. .. .. .. .. .. .. ..
23. Morphological and physiological variation among Aegilops squarrosa strains collected in Pakistan, Afghanistan and Iran
(KIHARA) .. .. .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
24. Karyotaxonomic studies in Poaceae, V. (TATEOKA) .. .. .. .. .. ..
25. Fertility variation due to "Gametic-Development Genes" inhybrid populations of rice (OKA)
26. Back-cross experiment between distantly related varieties of
rice-An interaction between maternal and gametic genotypes-(OKA) . . . . . . . . . . . . . . .. .. . . . . . . . . . . . . . . . . . . . . .. . . . .
27. A preliminary note on moist storage of dormant wild riceseeds (TAKAHASHI and OKA).. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

34
34

Cytology and genetics of Nicotiana and some other plants......
28. Cytogenetic studies on the genus Nicotiana X (TAKENAKA) .. . .
29. Notes on seedless Citrus species (FURUSATO and OHTA).. .. .. ..
30. Trifoliate seedlings in Citrus (FURUSATO, OHTA and ISHIBASHI)..
31. Heteroploid seedlings in Citrus (OHTA and FURUSATO)
32. Embryoculture in Citrus Natsudaidai (OHTA and FURUSATO).. ..

43
43
44
44
45
46

F.

32

36
37
39

40
42

CONTENTS
33.
34.
35.
36.

,

Study on fiber of tetraploid Luffa cylindrica Roem (FURUSATO)
A new type of bud variation in Citrus (FURUSATO)
Cytological studies in the Genus Euphorbia. II. Chromosome
numbers of twenty European species (SHIMOYAMA).. .. .. .. ..
Chromosome studies in Japanese Callitriche (SHIMOYAMA) ....

iii

48
48
49
50

G. Biochemical genetics of flowering plants
51
37. Leuco-anthocyanin in variable flowers of Camellia japonica(ENDO), .. .. .. .. .. 51
38. F 2 segregation of flower color in inter-varietal crosses of Swiss
giant pansy (ENDO)' .. .. .. .. .. .. .. .. .. .. .. .. 52
H. Population genetics and studies on quantitative characters. .. ..
39. Genetic analysis of a white leghorn closed flock apparently
plateaued for egg production (YAMADA) .. .. . . .. . . .. .. . . .. . .
40. Observation on natural populations of Formosan wild rice,particularly on variations in seed dormancy ~ TAKAHASHI and
OKA)
41. Genetical Studies on the cherry-red leaf in tobacco plants(SAKAI and IYAMA)
42. Genetic correlations of agronomic characters in rice (hAMA) ..
43. Migrating activity in inbred lines derived from two wild populations of Drosophila melanogaster (SAKAI, NARISE, ITo and
IYAMA), .. .. .. ..
.. .. .. .. .. .. .. .. ..
44. Variation in competitive ability among F 3 lines of the crossbetween an upland rice variety and" Red Rice" (lYAMA) ..
45. Genetic variability in wild populations of Oryza perennis andO. rufipogon (SAKAI and NARISE)
46. Heritability of grain shedding and other characters in rice
(SAKAI and NILES)53

I. Radiation genetics in animals .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
47. Radiation mutagenesis in the silkworm, I. Change in sensitivityof silkworm germcells to X-rays at different stage of gametogenesis (TAZUIl:A and ONIMARU), .. ..
48. Radiation mutagenesis in the silkworm, II. Mutation response
pattern of silkworm germcells to X-rays according to stage
and sex (TAZIMA and ONIMARU)
49. Radiation mutagenesis in the silkworm III. Comparison of Xray induced mutation rates in the silkworm with those of
Drosophila and mouse (TAZIMA),, .. .. .. .. ..
50. Tissue culture analysis of delayed lethal irradiation effect in

71

53

58
59
61

63
65
67
69

71

73

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CONTENTS

iv

51.

52.
53.

54.
55.
56.
57.

D. melanogaster (SUGAHARA and HORIKAWA)
Structure and function of pleiotropic genes in the silkworm, I.
An experiment to break up EEl, EKp and ENe genes into
separate components by X-rays (a preliminary note) (TAZIMA
and INAGAKI).
Silkworm larvae with extra-abdominal legs caused by a chromosome fragment (TSUJITA)..
Effects of chronic gamma irradiation on the breeding behaviorof mice at different developmental stages (SUGAHARA, TUTIKAWA, SUGIURA, and TANAKA).
Biochemical studies on the radioprotective effect of cysteamine(YAMAMOTO, NAWA and SUGAWARA)..
Melanotic tumors induced in Drosophila melanogaster by ionizing irradiation (TAIRA and MORITA)..
Differential sensitivity to X-ray irradiation of male and female
day-old chicks (KAWAHARA)..
A new spacing in the wide angle X-ray diffraction pattern ofdried striated muscle (SUGAHARA, OZAWA and UEZUMI).

J. Radiation genetics in plants
58.
59.
60.
61.
62.

63.
64.
65.
66.
67.
68.

.
Effect of X- and r-irradiation on chromosome aberrations in
Vicia faba (NEZU and MATSUMURA)..
Effects of temperature and irradiation time upon mutations
induced by radiations (MATSUMURA)..
Crosses between various X-ray induced recessive mutants inwheat (FUJII).
Radiosensitivity in plants (FuJII)..
Radiosensitivity of several commercial varieties and usefulmutants induced by X-rays in tobacco (MATSUMURA and
FUJII).
Induction of bud-sports by X- and r-rays (MATSUMURA and
FUJII).
Induced polygenic mutations in rice (OKA and HAYASHI).
Effects of r-radiations upon germination and growth of rice
plants (OHTA, KAWASHIMA and MATSUMURA).
Radiosensitivity and mutants in barley induced by irradiation(MATSUMURA and FUJII).
Effect of AET upon radiosensitivity of wheat seeds (MATSUMURA and KAWASHIMA)..
Diffusion of radioisotopes in soaked wheat seeds (KONDO, YOKOTA
and KAWASHIMA)
.

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76
77

79
81
82
84
85
87
87
88
90
91

93
94
95
96
97
97
98

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CONTENTSv

K.

Genetics of microorganisms
101
Transductional analysis of monophasic types of Salmonella (lINO
and LEDERBERG)
101
L. Mathematical genetics
102
102
70. Chance of fixation of mutant genes (KIMURA)
103
71. Effect of natural selection in a finite population (KIMURA)
72. Zygotic frequencies in a partially self-fertilizing population(KIMURA), . . .. . . .. 104
73. Overdominant genes in an partially self-fertilizing population(KIMURA), . . .. . . .. 105
74. On the decrease of fitness due to deviation from optimum inquantitative characters (KIMURA)
106
69.

M. Technical note
,
,
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107
75. Dose-rate distribution in the 60CO gamma-ray room (KONDO and
MATSUMURA)
107
76. Temperature dependence of the window glass dosimeter (KONDO) 110
112
Books and papers published in 1957 by staff members

•tGENERAL STATEMENT
In 1956 we have been very busy owing to the International Genetics
Symposia in Tokyo and Kyoto. This year we could return to our normal
laboratory life. Thanks to the kind support by The Rockefeller Foundation, we have started two new projects, one on the origin of cultivatedrice and the other on the effects of radiation on animals. One of themain objects of rice researches is to collect wild species and cultivatedvarieties of Oryza all the world over, so that we can reexamine the classification of the whole genus which may sheet light on the origin of rice.
The first collecting party headed by Dr. OKA was sent to Thailand,
India and Ceylon, where they have succeeded in collecting many seeds
and specimens of wild as well as cultivated rice. To our delight our projects have been received enthusioustically by many colleagues abroad,
who showed admirable cooperation spirit sending us many seeds. So far
we could obtain 16 species of Oryza. We are now confident that the remaining some 10 species will be collected in near future.
Studies on animal genetics started at the end of this year. We will
refrain from reporting until next year.
A group of our members initiated a study on the genetics and chemistry of bitter substances of Citrullus colocynthis. So far a bitter substance
was isolated which is called Citbittol. Taste blindness for the bitter substances was sporadically found among the populations near Misima andYokohama.
We had 3 visiting guest workers in Misima. Mr. RAJAN from New
Delhi stayed 8 months and studied the effects of X-rays on self-incompatibility of Brassica in the Laboratory of Dr. MATSUMURA. Prof. CROW worked with Dr. KIMURA during the summer. Prof. WARID has visited us in
the summer to get informations from various departments of the Institute.
However his main interests were concentrated in the methods of plant
breeding in Dr. SAKAI'S Laboratory.
Both Dr. YOSIDA and Dr. OSHIMA went to America for one year inorder to study in their respective fields. Dr. YOSIDA is working on cytology of tumor cells in Children's Cancer Research Laboratory in Boston.'
Dr. OSHIMA, a Drosophila specialist, is in the Carnegie Institution, Cold
Spring Harbor.
Prior to his trip to India, Dr. OKA went to U. S. A., Philippines andFormosa for 3 months and met specialists in rice plants. Dr. TAZIMA
went to India to visit sericultural experiment stations by the invitationof Ministry of Agriculture of India. He travelled up to Kashimir and2

ABSTRACTS OF DIARY FOR 1957

Assam in the North, and to Madras and Coimbatore in the South.
The first summer seminar on genetics for the senior high school teachers was held this summer. 72 teachers from 43 prefectures weregathered. The object of this seminar was to give sound informations on
genetic principles and to demonstrate the segregation and linkage ofgenes using plant and animal materials. The lectures on recent advancesin genetics were given by the members of the Institute. The seminar
was received favorably by the participants.

ABSTRACTS OF DIARY FOR 1957
25. 54th meeting of Misima Geneticists' Club.
4. Meeting of Tobacco Research Workers.
10. 8th series of Public Lectures on Genetics.
24. 16th meeting of Board of Councillors.
Mar. 13. Meeting of Tobacco Research Workers.
14. 55th meeting of Misima Geneticists' Club.
30. The laboratory for Microbiology and Tissue Culture, underconstruction since last year, was completed.
Apr. 25. 56th meeting of Misima Geneticists' Club.
May
1. Grant from The Rockefeller Foundation for the study on theorigin of cultivated rice was received.
4. 17th meeting of Biological Symposia.
21. 18th meeting of Biological Symposia.
23. Board meeting of Association for the Propagation of the Knowledge of Genetics.
25. 57th meeting of Misima Geneticists' Club.
June 7. 58th meeting of Misima Geneticists' Club.
14. 59th meeting of Misima Geneticists' Club.
21. Board meeting of Japan Association of Poultry Genetics.
22. General meeting of Japan Association of Poultry Genetics.
July 19. 60th meeting of Misima Geneticists' Club.
25-27. 1st series of Summer Seminar on Genetics
29. 61st meeting of Misima Geneticists' Club.
Aug. 6. 19th meeting of Biological Symposia.
10. Meeting of Editorial Board of "The Heredity" (Iden).
Sep. 27. 62nd meeting of Misima Geneticists' Club.
Oct. 18. 63rd meeting of Misima Geneticists' Club.
1. Grant from The Rockefeller Foundation for the study on theDec.
genetic effects of radiation in animals was received.
Jan.
Feb.

STAFF

3

STAFF
Director
Hitoshi KIHARA, D. Sc.

Members1.

Department of Morphological Genetics:
Yataro TAZIMA, D. Ag., Head of Department
Motoo KIMURA, Ph. D., D. Sc.
Kimiharu ONIMARU

2.

Department of Cytogenetics:
Yo T AKENAKA, D. Sc., Head of Department
Tosihide H. YOSIDA, D. Sc., Assistant Head
Tsuguo TATEOKA, and Kiyoshi TUTIKAWA

3.

Department of Physiological Genetics:
Chozo OSHIMA, D. Sc., Head of Department
Hitoshi KIHARA, D. Sc.
Toshifumi TAIRA, and Sadao SAKAMOTO

4.

Department of Biochemical Genetics:
Mitsuo TSUJITA, D. Ag., Head of Department
Yoshito OGAWA, M. D., Saburo NAWA, Bungo SAKAGUCHI, Seizo TSUDA,
D. Ag.
Toru ENDO, and Tetsuo IrNO (on leave of absence)5.

DePartment of Applied Genetics:
Kan-Ichi SAKAI, D. Ag" Head of Department
Hiko-Ichi OKA, D. Ag., Assistant Head
Yukio YAMADA, Akira MIYAZAWA, and Takatada KAWAHARA

6.

DeDartment of Induced Mutation:
Seiji MATSUMURA, D. Ag., Head of Department
Tsutomu SUGAHARA, M. D., Assistant Head
Sohei KONDO, Taro FUJII, and Yoshihiko SUGIURA.

Part-time Staff and Research Associate
Kan OGUMA, D. Ag., Ex-Director, Emeritus Professor of Hokkaido Univ.
Yoshimaro TANAKA, D. Ag., Emeritus Prof. of Kyushu Univ.
Taku KOMAI, D. Sc., Emeritus Prof. of Kyoto Univ.
Yoshinari KUWADA, D. Sc., Emeritus Prof. of Kyoto Univ.
Sajiro MAKINO, D. Sc., Professor of Hokkaido Univ.

4

STAFF

Hideo ETO, M. D., Assistant Professor of Tokyo Univ.
Kazuo FURUSATO,
Flora A. LILIENFELD, Ph. D.

Department of Administration
Kan-Ichi OTOFUJI, Head of Department
Sumiyoshi SUGIO, Head of General Affairs Section
Masao MIYAZAWA, Head of Finance Section
Naomi MATSUBARA, Hiroko NAKANO, and Junzo KADOWAKI
Clerks, Typists, Telephone operators, Chauffer, Field laborers andJanitors
24

Misima Branch of Hatano Tobacco Experiment Station
Masao TANAKA, Head
Flora A. LILIENFELD

Seiji IMAI
Assistants..... .4

Whole- Japan Association of Poultry Genetics
Hitoshi KIHARA, President
Kan-Ichi SAKAI, Vice-president

Association for Propagation of the Knowledge of Genetics
Hitoshi KIHARA, President
Yo T AKENAKA, Managing Director
Seiji MATSUMURA, Managing Director

COUNCIL
Yo K. OKADA, Director of National Science Museum, Chairman
Seiji KAYA, Professor of Tokyo University, Vice-Chairman
Sukehiro HIGUCHI, Director of National Institute of Radiological Sciences
Kan OGUMA, Emeritus Professor of Hokkaido University
Seizo KATSUNUMA, President of Nagoya University
Yohichi ASAMI, Emeritus Professor of Tokyo University
Toshitaro MORINAGA, Director of National Institute of Agricultural Sciences
Hideo MATSUKUMA, President of Japan Monopoly Corporation
Toshio SAITO, Governor of Sizuoka Prefecture
Yakichi NOGUCHI, Professor of Tokyo University
Riichi KAWAKAMI, Head of Department of Biometry, National Institute of
Public Health

RESEARCH PROGRAM FOR 1957

5

Kin-ichiro SAKAGUCHI, Professor of Tokyo Uuniversity
Masatada TERADA, Professor of Tokyo Jikeikai Medical College
Takeo SAKATA, President of T. Sakata Company,
Ichiro ISHIKAWA, Commissioner of Atomic Energy Commission
Daigoro MORIWAKI, Professor of Tokyo Metropolitan University

RESEARCH PROGRAM FOR 1957
Department of Morphological Genetics
Genetic analysis and linkage study in silkworm (TAZIMA)
Studies on a food preference mutant in silkworm (TAZIMA)
Structure and function of pleiotropic genes in silkworm (TAZIMA)
Genetic effect of radiation upon silkworm (TAZIMA)
Theoretical studies of population genetics (KIMURA)

Department of Cytogenetics
Cytology and genetics of tumors (YOSIDA)
Determination of sex and sex-chromosomes in animals (YOSIDA)
Cytogenetics with tissue method (YOSIDA)
Determination and differentiation of sex in plants (TAKENAKA)
Induction of abnormal mitosis and inhibitation of growth by substances
extracted from certain plants (TAKENAKA)•
Interspecifichybrization in Nicotiana (TAKENAKA, FURUSATO & LILIENFELD)
Genetics of Pharbitis nil (TAKENAKA et. al.)
Karyo-systematic studies in Poaceae (TATEOKA)

Department of Physiological Genetics
Genetical analyses on insecticide-resistance of Drosophila. (OSHIMA)
Genetical and biochemical studies on the metabolisms relating to the procedure of the eye pigment-formation in Drosophila (OSHIMA & TAIRA)
Fundamental studies on the effectiveness of ionizing irradiation on themetabolic processes in Drosophila (OSHIMA & TAIRA)
Developmental genetic studies of tissue-cultured eye discs of Drosophilamelanogaster. (OSHIMA)
Studies on the origin of wheat (KIHARA)
Studies on substitution of nucleus in what (KIHARA)
Studies on Agropyron (MATSUMURA & SAKAMOTO)

6

RESEARCH PROGRAM FOR 1957

Physiological genetics of the reaction of Japanese morning glory to daylength (SAKAMOTO)

Department of Biochemical Genetics
Biochemical genetics of insects and microorganism (TSUJITA, NAWA &SAKAGUCHI)
Embryological and biochemical studies of silkworm (TSU,JlTA & SAKAGUCHI)
Biochemical studies of some mutants in silkworm and Drosophila (NAWA& TAIRA)Biochemistry of the mechanism underlying variations in flower colors inplants (ENDO)
Biochemical studies on the differentiation of muscle protein in animals(OGAWA et al.)Biochemical studies on the mechanism of cell division in animals (OGAWA
et al.)
Biochemistry of the bitter substance in Citrullus colocynthis (OGAWA et. al).

Department of Applied Genetics
Fundamental studies on breeding and genetics in poultry (YAMADA &KAWAHARA)
Comparative Studies on poultry breeding using experimental animals(YAMADA & KAWAHARA)
Studies on polygenic inheritance (SAKAI et al.)Studies on competition between genotypes in plants (SAKAI et al.)Population-genetic studies of "Red-Rice" growing among upland rice
(SAKAI et al.)Studies on the technique of breeding in plants (SAKAI)
Genetic studies on "Cherry-red leaf" in tobacco plants (SAKAI et al.)Genetic studies on the migration activity in Drosophila (SAKAI et al.)Polyploidy and sterility in fruit plants (FURUSATO & MIYAeAwA)
Genetics of bitter-substance in Citrullus (FURUSATO)
Analysis of genes responsible for hybrid sterility and hybrid break-down
in rice (OKA)
Genetic studies of some physiological and agronomic characters in rice
(OKA)

Department of Induced Mutation
Measurement of X-, S- and r-ray dosage for induced mutations (KONDO)
Relation between the quality of radiations and mutations (MATSUMURA,
NEZU & KONDO)RESEARCH PROGRAM FOR 1957

7

Radiation genetics of wheat and barley (MATSUMURA & FUJII)
Studies on chlorophyll mutations induced by radiation (FuJII)
Mutation induced by irradiation of tobacco plants (MATSUMURA & FUJII)
Radiation genetics and its practical application (MATSUMURA et al.)Studies on radiation-induced mutations in mice (SUGAHARA et al.)Studies on the physicochemical mechanisms of radiation effects on livingorganisms (SUGAHARA et al.)Studies on radiation protections in animals (SUGAHARA et al.)
X-ray diffraction studies on muscle (SUGAHARA et al.)Improvement of sugar beets by means of induced triploidy (MATSUMURA& NEZU)
Research Students and Research Items
Hisataka ITO: Population genetics in animal breeding
Toshiteru MORITA: Biochemical genetics of amino acids and pterine
derivatives.
Yoshiyuki AMANO: Effects of antibiotics on cell division.
Tadao HAMADA: Experimental studies on the incidence of leucemia inmice.
Toshihide TABATA: Tissue culture studies on human chromosomes.
Sakinya FUJITA: Histological studies on genetic anomalies in mice.
liro SUZUKI: Studies on chromosomes of normal ciliar cells.
Hiroyuki HIRUMI: Developmental genetics by tissue culture.
Masakatsu HORIKAWA: Tissue culture and radiation.
Shinya IYAMA: Population genetics in crops.
Hiroko IKEDA: Statistical treatment in medicine.
Narito TAKAHASHI: Population genetics in rice.
Takashi OTA: Studies on plant improvement by radiation.
Yoichi YOKOTA: Radiation dosimetry.
Shoji KAWASHIMA: Applications of radio-isotopes in agriculture.
Masaaki ONOUE: Radiation genetics in mice.
Goro YAMAMOTO: Effects of radiation on nucleic acid metabolism.
Tetsuaki HASHIMOTO: Radiation biology in mice.

8

FOREING VISITORS IN 1957

FOREIGN VISITORS IN 1957
Jan. 29. Dr. R. F. CHANDLER. Jr. (The Rockefeller, Foundation).
Feb. 1 to Sept. 20. Mr. S. S. RAJAN (Indian Institute of Agriculture). Did
research work supported by the Colombo Plan.
May 1. Dr. R. K. ANDERSON (The Rockefeller Foundation).
3. Dr. W. A. WARID (University of Cairo, Egypt).
21. Dr. K. C. ATWOOD (Oak Ridge National Laboratory). Gave lectureon "Lethal mutation in Neurospora"
July 1 to Sept. 10. Prof. J. F. CROW (University of Wisconsin, U.S.A.).
Colaborated with Dr. KIMURA in a joint work on population
genetics. Gave lecture on "Genetic load of a population".
July 1 to 31. Assistant Prof. W. F. WARID (University of Cairo, Egypt).
Studied cytogenetics of vegetable. Gave lecture on "Present
status of plant breeding in Egypt"
July 19. Dr. H. L. EVERETT (Cornell University, U.S.A.).
24. Mr. D. M. SUMMENER (American Consul-General, Yokohama) andMr. M. E. LEE (Director of American Cultural Center).
26. Mr. and Mrs. E. WILSON (Chairman of the committee for Pasadena-Misima city affiliation).
Aug. 15. Prof. Abbas ARRUSHDI (University College of Arts & Sciences
Baghdad, Iraq).
Oct. 31. Dr. S. A. HOLMBERG (Algot Holmberg & Sons Ltd., Sweden).
Dec. 9. Prof. A. T. SANYAL (State College of Agriculture, Calcutta, India).

RESEARCHES CARRIED OUT IN 1957
A.

1.

GENETICS, BIOCHEMISTRY ANDCYTOLOGY OF INSECTS

Inheritance of a New Mutant" brown-3" of the silkworm(By Yataro TAZIMA)Light brown eggs are occasionally discovered in certain current commercial silkworm races of Chinese origin. It has been elucidated thatthe light brown egg is inherited together with transparent larval skin andblack eye color and that all of those traits are not separable but are controlled by one gene.
When light brown is crossed to normal, a marked difference is observed in the color of F I eggs according to the direction of the cross, due tothe after-effect of maternal genes. Among the F 2 eggs, i.e. eggs laid by
F I females, two different cases were observed. Usually segregation does
not occur within the same lot of the F 2 eggs, but in some instances segregation is clearly observed. Hence, the mode of inheritance of light brown
did not seem to be simple. Under certain circumstances it behaves as if
it were similar to bi and in other cases it behaves exactly as b2 • Bothbi and b2 genes are located on the sixth chromosome. Linkage test
between light brown and Ekp of the sixth chromosome indicated thatthey are independent.
A particular interaction was observed between light brown and some
egg color genes of the tenth chromosome. When light brown is crossed
to WI female, characteristic white eggs, which are usually expected inthis cross, become pigmented to an appreciable degree so that maternal
inheritance can hardly be ascertained. On the contrary, when light brown
females are crossed to WO l males, the resulting F I eggs remain mostly
white. By linkage test between light brown and fl (wingless, located on
the tenth chromosome), it was revealed that light brown is located on thetenth chromosome. Thus a responsible gene for light brown was foundto be different from bi and b2 and it was named brown-3 (b 3 ).
Three loci, i.e. WI, W2 and W3, have already been established on thetenth chromosome. It has also been known that crossing-over could be
detectable with small frequency between W3 and WO l , although both had
been reported to be allelic (KIKKAWA '47). The gene b3 is neither allelic
to WI nor to W2, but seems to be pseudo-allelic to W3. In combination

RESEARCHES CARRIED OUT IN 1957

10with W3, b3 hinders the transmission of 3-0H-kynurenine from egg
plasm to serosa cells, where it would be consumed in pigment formation.
This suggests that b3 and W3 might possess at least a part in common
such as a subunit gene within their gene complex.
Though no decisive proof has been obtained yet, I am inclined to assume that WO l belongs to the same group with b3 and W3. The hindrance
hypothesis for the transmission of 3--QH-kynurenine from egg plasm toserosa cells may well explain the occurrence of the two different segregation types in F 2 as stated above by assuming that the degree of hindrance varies with the environmental conditions.

2. Studies on a Food Preference Mutant in the Silkworm
VIII.

Selection for Higher Incidence of Non-preference
Individuals and their Genetic Analysis(By Yataro TAZIMA and Isamu MACHIDA)

Selection for higher incidence of non-preference individuals has been
continued up to the tenth generation in an X-ray induced mutant strainof the silkworm by beet-feeding test (TAZIMA '54). The results show aconsiderable selection effect, and the percentage of incidence of non-preference individuals increased from 0.15% in F I to 50.5% in FlO. In themeantime selection for higher hatchability has also been simultaneously
carried out in the same strain but it proved to be ineffective. In thisstrain hatching percentage has never exceeded 80% owing to a large
number of dead eggs in balance. This indicates that a lethal gene whichacts during embryonic stages is linked to the non-preference gene.
Crossing experiments were carried out in order to analyse the genetic
behavior of the non-preference character. The non-preference individualsof the F 9 generation were crossed to normal. In F I offspring hatchability
was recovered up to the level of the normal strain but incidence of nonpreference individuals decreased to 28.9%, i.e. approximately to half thelevel of FlO generation (50.5%). F 2 was obtained by sib-mating of nonpreference F I individuals; lethality appeared again resulting in 67.8%
hatchability, and the incidence of the non-preference individuals was
30.3%, which was slightly higher than in the preceding generation. The
non-preference individuals of F 2 were crossed again to normals. Incidenceof the non-preference individuals observed was 9.6 and 9.3% in the offspring of non-preference if- x normal '2)" and 5.0, 4.9 and 5.9% in theprogeny of normal if- x non-preference '2)".
The above results may well be explained by postulating that there ex-,

RESEARCHES CARRIED OUT IN 195711

ists a dominant main gene and several pairs of dominant modifiers thatcollaborate with the former in the manifestation of the non-preference
character and that the main gene is accompanied by a close linked lethal
gene. It was also noted that the cytoplasm might have some effect upon
the expression of the main gene.

3.

The Resistance of Strains of D. melanogasterto DDT and Dieldrin(By Chozo OSHIMA)

The strains used in this experiment were started from different females
captured in a single collection from each of several natural populations
in Europe, Africa, Japan and United States. These strains have been
cultured by mass-mating for about four years at Cold Spring Harbor andin Japan.
Forty female flies 4 or 5 days old, grown under optimal conditions, were
exposed to DDT, Dieldrin or control test papers in small vials. The test
papers were prepared by the WHO in Geneva. After exposure, flies
were transferred into a fresh vial containing a wet paper and the numberof dead flies was observed at definite intervals. Flies began to die bydesiccation and starvation 25 hours after exposure to control paper.
The levels of DDT-resistance in 18 strains of 10 localities were decided
on the basis of percentages of accumulated mortalities 25 hours after exposure to 4% DDT test paper for 5, 10 or 15 hours. On the other hand,
the levels of Dieldrin resistance in 11 strains of 5 localities were determined by percentages of accumulated mortalities 25 hours after exposure
to 0.4 or 0.8% DL test paper for 0.5, 1.0, 2.0 or 3.5 hours. The several
points showing each total average mortality of these strains at several
time doses are nearly linear by arranged on normal logarithmic section
paper. For this reason, resistance to both insecticides appears to benormally distributed in populations.
Eighteen strains were classified into three groups, non-resistant, slightly resistant, and resistant to DDT, and the numbers of dead flies at 25
hours after exposure to 4% DDT test paper for 15 hours were analyzed
statistically as to locality and strain. Highly significant differences in
DDT-resistance between localities and between strains were found; thelatter were larger than the former but not significantly so. Eleven strains
were also classified into three groups, non-resistant, slightly resistant, andresistant to Dieldrin and the numbers of dead flies at 25 hours after exposure to 0.8% DL test paper for a half hour were analyzed statisticallyas to locality and strain. Highly significant differences in DL-resistanceRESFARCHES CARRIED OUT IN 1957

12

between localities and between strains were found again but in this casethe former were significantly larger than the latter.
These results indicate conclusively that any local population is extremely heterogeneous in DDT and Dieldrin resistance; locality-strain differencesin two cases may be attributed to different use of the insecticides.

4. Genetical and Biochemical Studies on Pterine Oxidation
in Drosophila melanogaster(By Saburo NAWA, Toshifumi TAIRA and Bungo SAKAGUCHI)

The Drosophila homogenate is capable of oxidizing both xanthine and2-amino-4-hydroxypteridine (AHP). It is likely that the pterine dehydrogenase in Drosophila is a dehydrogenase, not an oxidase, further that DPN
is a more effective hydrogen acceptor than methylene blue (MB), and alsothat the conversion of AHP to isoxanthopterin is carried out by pterine
dehydrogenase in the presence of DPN in vivo. The enzyme preparation
was made from pupae of the wild type of D. melanogaster. The freshly
prepared supernatant produces a considerable amount of isoxanthopterinfrom AHP in the absence of any external (exogenous) hydrogen acceptor.
However, when the dialyzed enzyme or aged supernatant is used, noappreciable production of isoxanthopterin is observed without an external
acceptor. The activity of different preparations in the presence of various
acceptors is shown in Table 1.
Table 1.

Effect of electron acceptor on the oxidation of AHP in
preparations from Drosophila.
Preparation

Acceptor
fresh

aged

dialyzed

0.55*
0.9
0.9

0.08
0.58
0.6
0.1

0.02
0.7
0.75
0.02

~ _ - - - -

Water
MB
DPN
Cytochrome c* Activity was expressed in fJ. moles of isoxanthopterin produced per gram
whole pupae (wet weight) per hr (pH 8.0).
The activity of pterine dehydrogenase in the presence of DPN has beenfound to be about equal to or rather higher than that of ME. It seems
that DPN naturally present in Drosophila acts as an hydrogen accetor.
Under aerobic conditions, isoxanthopterin of much higher concentration
is produced than that of DPN added. The relation is shown in Fig. 1.

RESEARCHES CARRIED OUT IN 1957

13

This may be because pterine dehydrogenase is linked to the DPNH oxidase
system. The pH optimum of DPNH oxidase in Drosophila is in the neighborhood of 6.5. The ratio of the reaction rate with DPN and with MB
as an acceptOl is very
different at various pH
30
levels. This may be due
Initial DPN concentratwn: /0-5 M
to the participation of ..,DPNH oxidase in thereaction. The action of
2.0
the enzyme for pterine
oxidation in Drosophila isinfluenced by the concentration of DPN andthe rate of DPNH reoxidation. For example,
it has been known thatthe mutant rosy lacks
10
20
30
Time in minutes
pterinedehydrogenase.
The ratio of the reaction
Fig. 1. The effect of the re-oxidation of DPNH
on the oxidation of AHP.
rate in the presence of
DPN and that of MB as
an acceptor varied considerably in some different mutants. This may be
due to the difference of the rate of DPNH oxidation. Further experimentsin this line are in progress.

~

5.

Purine Catabolism in D. melanogaster(By Toshiteru MORITA and Chozo OSHIMA)

It is well-known facts that the rosy" (ry") of the eye color mutants in
D. melanogaster does not contain isoxanthopterin which occurs widely in
Drosophila, and that the enzyme prepared from Drosophila can catalyze
the oxidation of 2-amino-4-hydroxypteridine (AHP) to isoxanthopterin as
well as hypoxanthine and xanthine to uric acid. The enzyme, therefore,
has been hitherto called xanthine oxidase or pterine oxidase, but it is true
dehydroganase because of requirring an electron acceptor. As an electron
acceptor, diphosphopyridine nucleotide (DPN) seems to be more efficientin vivo, but it is rather less effective than methylene blue in case of
purine-catabolism.
An eye color mutant, rosy (ry), does not contain a trace of isoxanthopterin as likely ry", and it accumulates a larger amount of hypoxanthineand xanthine in stead of uric acid at pupal stage as following table.

14

RESEARCHES CARRIED OUT IN 1957
Table 1.

Pteridines and purines found in D. melarwgaster strains.
Oregon-R

ry
A

AHP
Isoxanthopterin
Hypoxanthine, Xanthine
Uric Acid

LarvaePupae

Adults

Larvae

±
±
±
±

+

+
+

±

±

±

*±
+

+

Pupae Adults

*
*

+
+

Furthermore, the ry strain does not show any activity of xanthine dehydrogenase, and it seeems to be due to a lack of the enzyme. The same
phenomenon are found in such double recessive mutants homozygous forry, as v; ry, cn; ry, bw; ry and se; ry. From these results, it seems
that the ry gene relates to both purine- and pterine-catabolisms as following scheme.
AHP ------[-----> Isoxanthopterine----- ry ------1
Hypoxanthine ----1--> Xanthine ---,---> Uric Acid

6.

Dopa Oxidase in the Eye-color Mutants of Drosophila melanogaster(By Toshifumi TAIRA and Saburo NAWA)

In regard to the activation of tyrosinase in Drosophila, some findings
have been reported. A comparative analysis on the activity of Dopa oxidase of the eye-color mutants in Drosophila was hardly ever before carriedout. Such eye-color mutants having yellow pigment as se or Hn r3 strain
were used to analyze the Dopa oxidase activity in comparison with thewild type strain. As a rule, Dopa oxidase is completely activated when
the homogenate of wild type prepupae was kept at oDe in an ice-bath forone hour. Whereas, Dopa oxidase activity in the activated homogenate of
se was much lower than that of wild type. This may be due to someinhibitation-mechanism interfering with the process of activating the enzyme.
It seems that the inhibition-mechanism is probably related to the processof yellow pigment formation.
7.

Formation of Drosophila Eye Pigments(By Toshifumi TAIRA and Saburo NAWA)

It has been known that the wild type eye-color of Drosophila melanogaster is due to the presence of two pigments, the red and the brown.

'f

15

RESEARCHES CARRIED OUT IN 1957

It is seemed to be of particular interest to study the behavior of pteridine,since the red and yellow pigments are its derivative. The analysis of
pigments and pterines was carried out by paper chromatography. The
results are summarized in the following tables.
Table 1. Phenotypic effects of gene-combinations.
=========~'~~-~~~~-~~~~~-~-~--

Genotype

Phenotype

Genotype

Phenotypew; enno colorno colorno colorno colorno colorv; bw

no colorno colorbw typebw typebw type

w; bw
w;

8e

w; Hn r3w; ry

en; bw
bw; Bebw; Hn r3
bw; eaGenotype

Phenotypev; el

cl typeBe typeHn r3 typelight red
light redv; Be

en; Hn r3v; ryen; eaThe mutant genes se, cl, and Hn-r3 suppress the formation of the red
pigment and accumulate the yellow pigment in eyes. Therefore, the red
pigment probably derived from the yellow pigment. In the body of mutant
Table 2.

Relative amounts of pigments and pterines in single andcompound mutants (7 day-old).
I"

Mutants

J

-_.

v

Or-2

en

ea

en;ea

cl

v;cl

-------------------~Eye

Red pigment

-Itt

-Itt

,I Yellow pigment

±
±

±
±

+

_. ~I_~~;*nthoPterin
Body

_..

+

+

+

±

±
±

±
±

+

+

+

+

*± *±
*± *±

*±

; Yellow pigment

±

±

±

±

±

Isoxanthopterin
AHP*

li-ltt±

-Itt

*±

*±

-Itt

-Itt

±

±

±

ry

bw

w

II

±

+

--------------_.-.-_ .. _ - - - -Mutants

Eye

Red pigment
Yellow pigment
Isoxanthopterin
AHP*

I Be

'v;se

T~-- ±

I; ;----:-------Body

Yellow pigment
Isoxanthopterin
AHP** AHP:

Hn r3 en; Hn r3

+

+

+

+

+

±

±

±

+

+

*

±

±

-Itt

*+

*+

±

-Itt
±

±

±

±

-Itt

2-amino-4-hydroxypteridine

16

RESEARCHES CARRIED OUT IN 1957

Hn ra , accumulation of yellow pigment and decrease of isoxanthopterin are
observed simultaneously. This suggests that the normal metabolism of
pteridine is blocked by the gene Hn ra and that the yellow pigment is aprecursor of isoxanthopterin. The mutant ry is not capable of oxidizing
AHP because of the absence of pterine dehydrogenase. However, it remains to be elucidated whether or not the low ability of the formation of
red or yellow pigment in ry is linked to the absence dehydrogenase.
8.

Genetic Determination of Tyrosinase and Protyrosinase in Bloodof Silkworm, Bombyx mori L.
(By Bungo SAKAGUCHI)

The mutant, used in this experiment, with respect to low tyrosinase
activity in the blood of larva was found recently as a spontaneous mutant
in green cocoon (Gc) inbred strain at this institute. The mutant was
temporarily designated as ty.
1. Genetics of ty.
Tyrosinase activity in the blood of F 1 hybrid between a wild type (N124)
and ty showed an intermediate value of both parents. From the resultsof backcross and F 2 segregation, it seems to be conI
trolled by a single ressecive
~,400
gene.
l \
wtld type
2. Activation of ty ty:'eyrosinase.
/\
I\
Blood collected from the9\I\
5th inster larvae was used for<:ithe analysis as a source of the----r/\~ .200I\
I\
eenzyme tyrosinase. Solid ammonium-sulfate was added to/\the blood to 30% saturation
I',
I\
and the precipitate which conI\I\\
tains the enzyme was centri,,,J
fuged down and dissolved ino L-CtJ-ln-'t-.4+.0c---'---.S!;;-O-----''--f60;;--..L---.7~O,.-'---;;80;;a half volume of distilled watertemperrzture (Oc)
per volume of the original
Fig. 1. Relations between enzyme activities
blood. The dissolved solution
and various heat-treatment.
was then dialyzed over night
against distilled water in the vacuum condition. The dialyzed solution
consisted of partially purified enzyme preparation was used as the enzyme
for the analysis of this experiment.

",

\

,I

\

RESEARCHES CARRIED OUT IN 1957

17

The tyrosinase activity was determined with photoelectric colorimatorusing filter of 471 ml-t. The reaction mixture contained 0..1 ml of enzyme
preparation, 0.5 ml of 0.01 M dopa solution and 4.1 ml of 0.1 M phosphatebuffer, pH 6.8. Readings were taken at 5 minute intervals.
The heat-activation experiments were carried out for 5 minutes at thetemperature ranging from 45°C to 80°C with 5°C intervals. These treatments were made prior to the determination of tyrosinase activity. The
results are given in Figure 1. As shown in the table, pretreatment of
55°C for 5 minutes gave the highest activity in both enzymes obtained

.600

~

1\
:~ \
I'

tI \\

;:::

tyrosinase

prolyrvsLnase:I \\'<t ,400

•

"
"

II
,I

II
"

\\
II

!I \\

I,If

,: ,I

l'l

i

!I

J I

.200

"

...

/
----'

,

"/ I

"

wLld type

o

ly

\\
"

\" \
\\

\\
\ I
'\

,

", .
"

\~ -

---..._--..."",.,. , .........' .........
o L~=t:=:::::L----,'-c-~:::.::S~S==4~o===:f...·---towith (NH4 ),SO.

Fig. 2.

Solubility of tyrosinase and protyrosinase in (NHj)::S04 solution.

from ty and wild type silkworms. The activation rate of ty enzyme wasmuch higher than the wild type enzyme under those conditions given.
In addition to the heat-activation, activations of enzyme tyrosinase weremade by such treatments with acetone, chloroform, ethyl-ether, ethyl-alcohol and temperature at ooe for 1 to 3 hours.
The enzyme tyrosinase activated by various treatments mentioned above
belongs to a catelogy of protyrosinase or latent phenola:3e found in eggsof Grasshopper Melanoplus differentialis (BODINE and ALLEN, 1938), DrosophivJ

18

RESEARCHES CARRIED OUT IN 1957(OHNISHI, 1954, HOROWITZ and FLING, 1955) and Broad-Bean (KENTEN, 1957).
It was found from these results that protyrosinase in the blood of t y wasmuch more accumulated than that of wild type. The accumulation of
protyrosinase was controlled by the ty gene.
3. Solubility in ammonium sulfate solution of tyrosinase and protyrosinase
contained in ty and wild type (+t1I).
Each 20 ml of blood of ty and +t y strains were separately collected,
and the same volume of 0.1 M phosphate buffer, pH 6.3 was added to theeach blood sample. Solid ammonium-sulfate was then added to the mixed
solution to give 5 to 60% saturated concentration. The precipitate wascentrifuged down and dissolved in same volume of distilled water pervolume of the original blood. The dissolved solution was used as thetyrosinase preparation. This preparation was heated at 55°C for 5 minutes
and used as protyrosinase in the following analysis.
Measurement of enzyme activity were carried out with photoelectriccolorimeter under the conditions described earlier. Results are graphicallysummarised in Figure 2. The greater parts of both tyrosinase containedin blood of ty and + ty strains were collected in the fraction of 20% satulated ammonium sulfate. It was found that the solubilities of tyrosinaseand protyrosinase proteins in ammonium-sulfate solution in both ty and+ ty strains are of distinctly similar in nature. It seems that the gene ty
controls the specificity of the structure of the enzyme protein or that ofactive site of the enzyme protein.

9.

Chromosomal Constitution of the Original Nl 2 Strain of the Silkworm(By Mitsuo TUJITA)U, N1 2 , oa and several other genes belonging to the 14th linkage groupof the silkworm form the pseudoallelic U-complex (or U region). The
gene oa is recessive, and larvae homozygous for oa exhibit a transparent
hypodermis. However, larvae with the genotype Nl 2 /oa have a transparent
hypodermis. This fact suggests that Nl 2 is due to a deletion involving theoa locus.
The genetic behavior found in the progeny of the cross between theoriginal Nl 2 strain and oa/oa indicates partly trisomic inheritance andpostulates the presence of a fragment of chromosome XIV. In the lightof this hypothesis the following diagram illustrates the genetic constitution of the original Nl 2 strain.
The cytological and genetical study of this strain will be continued.

RESEARCHES CARRIED OUT IN 1957

~
+NZ

19

Attachment of spindle fibres
2l--------'ieP

Chromosome fragment involving the + B1 2 segment

+011

~dk

NZ'2
[1_ _-+-=-1_

N12 is due to the deletion of the +N12 segment

Attachment of spindle fibres

Fig. 3.

Diagram of the genetic constitution of the original Nl 2 strain.

10. Studies on the Mitochondria in the Gland Cells of Malpighian
Tubules of the Silkworm Larvae by Means of Ultra-thin sections(By Mitsuo TSUJITA)

In my previous papers (1937, 1947), the morphology of mitochondria andand their relation to secretion in the Malpighian tubule cells of the silkworm larvae were described. In recent years I have continued to study
the mitochondria in the cells of Malpighian tubules by ultrathin sections.
Principal points of my observations are as follows:
The inner margin of gland cells is bordered with a brush border which
consists of cytoplasmic protrusions in which mitochondria with cristae-likestructure are observed.
At the beginnig of each instar mitochondria which increased their number in the cytoplasm migrate to the base of the inner margin of the glandcells and protrude into the gland lumen together with a small amount ofcytoplasm. Thus, the brush border is formed. Cytoplasmic protrusions containing mitochondria, sometimes show rod-, clavate- or tadpole-shape. At
the middle stage of each instar, except for the tubules of the outer layer
in the rectal wall, various phases were found which indicate that filamentous mitochondria migrate from the cell body to the base of the protoplasmic processes driving toward the preformed process border. In thus
manner, protoplasmic processes can be supplemented at the middle stageof each instar. It seems that in the winding as well as ascending parts,
often processes change into needle-- or rod-shaped cristal. And as statedin my previous paper (1937), in the later stage of each instar granular
mitochondria migrate into the lumen of the gland, where they disappear.

RESEARCHES CARRIED OUT IN 1957

20

Similar cytological structures can be observed in the goblet cells of the
mid-gut epithelium of the silkworm larvae. The goblet cell is occupied
for the most part by a large vacuole or goblet, which is flask-shaped.
The inner surface of the goblet is apparently bordered by a mitochondrial
layer embedded in an acidophile protoplasmic matrix, in which fibrous
mitochondria are arranged almost parallel to each other. It seems to bereasonable to assume that in this case the mitochondrial layer is specificform of brush border, a layer of cytoplasmic protrusions, in each of which
fibrous mitochondria are contained. The so-called" SUibchensaum" of thecylindrical cells is a typical form of brush border which consists of anumber of protoplasmic protrusions.
Recently, BEAMS, T AHMISIAN and DEVINE (1956) worked with electron
microscope on the cells of Malpighian tubules of the grasshopper paying
special attention to the behaviour of mitochondria in the gland cells. In
their paper they describe that filamentous mitochondria migrate from thecell body to the base of protoplasmic processes where they protrude into
the inner margin.

B.

11.

CYTOGENETICS, GENETICS, CYTOLOGY AND
BIOCHEMISTRY OF TUMORS

Two Chemicals Promoting Cell Divisions in the Yoshida Sarcoma(By

Yoshito OGAWA, Kenjiroo FUJIOKA and Yukihide ABE)

We found that two chemicals act as mitotic stimulants on animal cells.
One of them is Kinetin and the other is Na-Glucuronate.
Kinetin (6- Furfurylaminopurine) was isolated by MILLER et al. (1955, 1956)
from deoxyribonucleic acid from herring sperms as a stimulant of plant
cell mitosis. Its effects on the division of animal cells, however, was notwell known, though negative results have been reported by LETTRE et al.
(1956) using animal and human cells. We have investigated the effect ofKinetin on the mitotic activity of YOSHIDA sarcoma cells transplanted
to the Wister strain of rat. Injecting the solution of Kinetin into theabdomen (most effective dose is 0.15 mg. per 100 g. body weight), a significant increase in the frequency of mitotic figures was found in the tumor
cells after about 120 hours (significant at the 19,6 level), as shown in Fig.
1. (OGAWA, Y. et al.: Nature, 180: 985. 1957)
Na-Glucuronate is well known as a growth promoting substance inanimals. We have investigated the effect of Na-Glucuronate on the mitotic
activity of YOSHIDA sarcoma cells using the same method as in our experi-

..

21

RESEARCHES CARRIED OUT IN 1957

ment with Kinetin. When a solution of Na-Glucuronate was injected into
the abdomen of rats (most effective dose is 750 mg. per 100 g. body weight),
a significant increase in the frequency of mitotic figures was found in thetumor cells after 24 (significant at the 5% level) and 96 (significant at the
1% level) hours. But when injected into the muscle tissue, no significant
increase was found. This suggests that effect of Na-Glucuronate on sarcoma cells may be direct.
We can find no influence upon the frequency of mitotic stages in sarcoma cells either when Kinetin or Na-Glucuronate is injected.
Further, to examine the effects of promoting activity at a combined
treatment of the YOSHIDA sarcoma with both chemicals, cell division wasinvestigated using the same method as in the above experiments. At a
combined treatment, the promoting effect of Na-Glucuronate on sarcoma
cell divisions was not investigated 24 hours after transplantation. But
after 48 hours, we could not fined a statistically significant difference
between separate and combined treatments as shown in Fig. 1.

200

100
Contr.

//\

R.

(0.15mg.

.

\

\

\

\

'. \

-~-

10

d.

Fig. 1. Daily change in frequency of metaphasic cells (pathologicalfigures excluded) after injecting Kinetin, Na-gluculonate and bothchemicals combined into the abdomen of rat, 10,000 cells being
observed per day.

This result indicates that:
1) The effect of Na-Glucuronate which appears 24 hours after transplantation of sarcoma cells is due to a different mechanism from thatwhich acts 96 hours after transplantation.
2) The two chemicals do not help each other in promoting cell divisions
in the sarcoma 48 hours and later after transplantation of sarcom cells.

22

RESEARCHES CARRIED OUT IN 1957

12. Biochemical Similarity of Mitotic Stimulants in Normal andSarcoma Cells of Rat(By Yoshito OGAWA)
MALMGREN (1955) reported that the saline extract of mammary tumor ofmice, if injected into the abdomen of rat, produced a significant increaseof the mitotic index in the liver after 48 hours. In order to test if his
findings are limited to his materials or can be generally true, we investigated mitotic indexes in the host liver after subcutaneous transplantationof a spontaneous fibrosarcoma which had been obtained from a Waynepink-eyed yellow rat.
Four days after the transplantation, a significant increase of mitotic
index was observed in the host liver as shown in Table 1. It is surethat this was not due to our misjudgment of metastasis of transplanted
sarcoma in view of the following points:
1) The significant increase in mitotic index appeared before the growthof transplanted sarcoma and disappeared 7 days after transplantation.
2) This fibrosarcoma was only transplantable to the Wayn-pink-eyedyellow strain of rat.
Therefore, it may be generally recognized that some mitotic stimulant
produced by tumor tissue is effective in normal cells. (OGAWA, Y. et al.:Med. and Biol. 45: 110. 1957)
Is the mitotic stimulant produced in normal tissue also effective in sarcoma cells?
200

100

10

d.

Fig. 1. Daily change of frequency of metaphasic cells (pathologicalfigures excluded) after injecting saline extract of rat embryos into
the abdomen of rat, 10,000 cells were observed per day.

,

23

RESEARCHES CARRIED OUT IN 1957
Table 1.

Mitotic index in transplanted sarcoma and in host liver tissue
Mitotic Index inTransplanted
SarcomaDays after
Transplantation

Mitotic indexinHost Liver

Host~--------------;---------,---------i---------

4

6.7
3.3
25.0
10.0

Wister

8.3
13.3
0.0
3.3

Hybrid*

6.7
45.0
6.7
10.0
0.0

Wayne

-----1

7

23.3
20.0
58.3

0.0
0.0
0.0

14

0.0
80.0

0.0
13.3

21

85.0
36.7

16.7
1.7

28

40.0
6.7

8.3
0.0

I
I

Wayne

-

Control

0.0
1.7
0.0

Wister

0.0
0.0
0.0

Hybrid*

0.0
0.0
0.0
0.0
1.7

Wayne

- - - - - - - - - - - - - - - - - ---~----------'--* F 2 of Wister x Wayne
In order to answer this question, saline extract of rat embryo(Wister: 4 and 20 days old) was injected into the abdomen of the samestrain of rat (Wister: 2 month old) immediately after the transplantationof YOSHIDA sarcoma, and the daily change of frequency of metaphasic
cells was observed. As shown in Table 2, when saline extract from the

RESEARCHES CARRIED OUT IN 1957

24

tissue of 4 days old embryos was injected, a significant increase in thefrequency of mitotic figures was found in the tumor cells after 24 and96 hours (both significant at the 5% level). But when the extract fromthe tissue of 20 days old rat was used, no significant increase was found.
It may then be concluded that the mitotic stimulant produced by sarcoma
tissue is effective in normal cells and that produced by normal tissue iseffective in sarcoma cells.
This suggests that there is something in common between normal andsarcoma cells in producing, and responding to, mitotsis is stimulating substance.

13.

Idiogram of a New Subline of the Yoshida Sarcoma
and Some of its Properties(By Tosihide H. YOSIDA)

As stated in the previous report (YOSIDA 1957)*, tumor cells characterized
by a new chromosomal constitution were observed in the YOSHIDA sarcoma.
These cells had 40 chromosomes, including one strikingly large J- and twolarge V-elements, and they gradually increased in number in succeedingtransplant generations. At the twentieth transplant generation the majorityof tumor cells showed a new karyotype. Thus, a new subline was established in the course of serial transplantations.
Idiogramatic analysis discloses that these tumor cells have 15 rod-, 7 Jand 18 V-shaped chromosomes, among which two V's and one J are remarkably large in size. On the other hand, the original tumor cells contained 15 rod-, 8 J- and 17 V-chromosomes. On the basis of karyologicalanalysis of these tumor cells, it appears probable that the large J-shaped
element, which characterized the new subline, resulted from translocationof two long arms of the first and the sixth J-shaped elements. Furthermore, a comparison of numerical difference in the J- and V-shaped chromosomes of the two groups suggests that the short arms of two J-elementsof the original cell combine to form a small V-shaped element by meansof centromeric fusion.
Transplantability of this subline increased gradually, following the increase in number of tumor cells having the new karyotype. The hostrange for adaptability of new tumor cells was wider than that of the original tumor cells. The average life span, observed in 153 Misima-bred
rats bearing the new subline, was found to be 10.3 days. The average
life span of 21 Misima-bred rats bearing the stock of the Yoshida sarcoma
obtained from the Sasaki Medical Institute, Tokyo, was 13 days, while thatof 34 Misima-bred rats bearing another tumor strain from the Pharmaco-* YOSIDA, T. H. 1957.

Ann. Rep. Nat. Inst. Genet. Japan 7: 16-17.

RESEARCHES CARRIED OUT IN 195725

logical Research Foundation, Tokyo, was 12 days.
It may be concluded from these observations that the competitive abilityof the new tumor cells which developed by the chromosomal translocation,is superior to the original form.

14.

A Karyological Study on the Yoshida Sarcoma Cells Infiltrated
into Organs of a Tumor-bearing Rat(By Tosihide H. YOSIDA)

As mentioned in the foregoing paper (in this report) new tumor cells of
the YOSHIDA sarcoma are characterize1 by one large J- and two large Vshaped chromosomes in their chromosbme complex. These cells gradually
increased in number during the course of ensuing generations. At theninth transplant generation the percentage of the new tumor cells to theoriginal cells was 79.0 per cent in the ascites tumor, while at the tenthtransfer generation this percentage increased to 88.9 per cent.
The following experiment was carried out in order to investigate theinfiltrating ability of the new and the original tumor cells into organs ofa tumor-bearing rat at the ninth transplant generation. From this rat,
small amounts of the lung, the spleen and the liver were inserted separately into the peritoneal cavities of normal rats. In the event that tumor
cells would infiltrated into organs readily, implanting the minced organs
into normal hosts would serve as an inoculum, and give rise to a new
ascites tumor population.
The observations have disclosed that in the case of transplantation of
the liver, 90 per cent of tumor cells were of the new karyotype; while inthe case of the lung and spleen transplantatifJlls. only 33.3 and 2.6 per centof tumor cells had the new karyotype, respectively.
Based on the above investigations, it seems probable that the new tumor
cells display a higher degree of infiltration in the liver as compared tothat in the spleen and the lung.

15.

Karyological Study on the Normal Somatic and Malignant
Tumor Cells in the Human(By Tosihide H. YOSIDA and Toshihide TABATA)

The chromosomes of normal somatic cells in six human embryos andone adult were studied using squash technique-the material was pretreatedin hypotonic solution*. The chromosomes were opserved in twelve somatic cells which showed a clear metaphase condition (Figs. 1 and 2). Variation of the chromosome number in these cells is shown in the Table l.
As shown in the table the cells having 46 chromosomes were most frequent.

* YOSIDA, T. H. and T. ISHIHARA 1956.

La Kromosomo 29: 1005-1009.

RESEARCHES CARRIED OUT IN 1957

26

The chromosomes of tumor cells obtained from the material of the stomach cancer, the tongue cancer, the maxillary cancer which developed bymetastasis of the stomach cancer, and the HeLa cells cultured in vitro
were studied. The number of chromosomes in the stomach cancer cells
showed two peaks of variation, one of which is 60 to 70, and the other is
100 to 110. The tongue cancer cells contained from 40 to 50 chromosomes.
On the other hand, the chromosome number of the maxillary tumor cells

Figs. 1 and 2. Chromosomes of a male germ cell in a human embryo (2n=46).
Fig. 1 is a microphotograph, Fig. 2 is a drawing of the same cell as Fig. 1.
Table 1.

The variation of chromosome number in normal somatic
cells of the human.

No. of chroms.
No. of cells ~~ser_v_e_d--,I

43

44

45

o

O_~__7

46

47

48

Total

3

12

showed a variation from 30 to 40. It is readily noted that the majority ofthe original tumor cells in the stomach cancer has a hypotriploid and hypopentaploid chromosome constitution, whereas the tumor cells in the
maxillary tumor, which developed from the metastasis of the stomach canc,er, showed a hypodiploid chromosome constitution. It has been noted by
YOSHIDA (this report) that the infiltrating ability of the YOSHIDA sarcoma cells
into organs of a tumor-bearing rat varied according to the difference of
karyotypes in these tumor cells. It is suggested from this study that thechromosome constitution of the maxillary tumor cells differ from those of theoriginal stomach cancer on acount of selective adaptation of metastatic cells.
HeLa cells, grown in tissue culture, showed a variation from 80 to 90
chromosomes.

RESEARCHES CARRIED OUT IN 1957

16.

27

Change in the Transplantability of the MY-mouse Sarcoma in
Relation to t~e Change of the Chromosome Number.

(By Tosihide H. YOSIDA and Tadao HAMADA)

Studies on the transplantability of the MY-mouse sarcoma were carriedout by YOSIDA (1952)*, ISHIHARA and YOSIDA (1954)**, and YOSIDA and ISHIHARA (1955)***. According to their observations this tumor was transplanted successfully to the S, Sk, S4' D and D-103 strains of mice, whichwere inbred in this institute. On the other hand, this tumor failed to
grow in strains of mice such as C3H, DBA/2, A, SWR and Swiss Albino,
imported from the United States. Present studies on the transplantabilityof this sarcoma have revealed that the host range can be conspicuously
extended to include such mouse strains as D, D-103, S, S4' C57L, dba,
DBA/Ma, Swiss Albino, C57BL, dd, C58, SPS, A and C3H.
YOSIDA (1954)**** studied the chromosomes of this sarcoma and noted it
to contain about 40 chromosomes in number. It is interesting to note that aremarkable increase in the chromosome number was observed in this study.
The majority of cells have 80 chromosomes. The stem line, which ischaracterized by tetraploid cells, was derived from the diploid stem line.
These studies illustrate that change in the host range of transplantable
tumors are related to chromosome ploidy.

17. Effect of 8-azaguanine and its Related Compounds on
the Chromosomes of the Yoshida Sarcoma Cells(By Tosihide H. YOSIDA and Hiroyuki HIRUMI)
8-azaguanine which is said to be antagonistic to pyrimidine metabolism
causes the deformation of chromosomes in the YOSHIDA sarcoma cells.
Normal chromosomes of the YOSHIDA sarcoma are generally characterized
as rod, J- and V-shaped. By the intraperitoneal injection of 8-azaguanine,
the shape of chromosomes of the tumor cells was observed to change toa round shape.
In order to clarify the influence of 8-azaguanine on the chromosome deformation, seven chemicals structually related to 8-azaguanine were employed in this study. They are grouped by their chemical constitutions asfollows: (1) Compounds having triazolpyrimidine nucleus with a hydroxygroup (-OR) at the 6-position of the pyrimidine nucleus. (2) Compound* YOSIDA, T. H. 1952. Jour. Fac. Sci. Hokkaido Univ. Ser. VI. Zool. 11: 41-50.
** ISHIHARA, T. and T. H. YOSIDA 1954. Ann. Rep. Nat. Inst. Genet. Japan 4:
15-17.

*** YOSIDA, T. H. and T. ISHIHARA 1955. Ibid. 5: 23-24.
**** YOSIDA, T. H. 1954. Ibid. 4: 2.

28

RESEARCHES CARRIED OUT IN 1957

having triazolpyrimidine nucleus with a 6-oxygen, instead of the 6-hydroxygroup. (3) Chemical having triazolonly. (4) Chemicals having imidazolpyrimidine with a 6-hydroxy group on the pyrimidine nucleus. (5) Chemical having imidazolpyrimidine with 6-oxygen.
The experiments were carried out as follows: 0.5 cc of ascites tumor
was mixed with each compound in vitro, and immediately injected into
peritoneal cavity of normal W-strain rats. The ascites cells were examined
6, 24, 72 and 120 hours after injection by the usual squash technique.
Our observations revealed that the compounds of group 1 were most effective for chromosome deformation, showing as great degree as the 8azaguanine. On the other hand, compounds of groups 2, 3 and 4 showedabout half the effect of 8-azaguanine. No effect was found in the compound of group 5. These observations suggest that the chromosomal deformation is caused by compounds containing a certain chemical nucleus
and/or a specific functional group, such as the triazol nucleus and the 6hydroxy group.

18. Effect of Chemical Anions on the Spreading and Splittingof Animal Chromosomes(By Tosihide H. YOSIDA and Tosihide TABATA)
By using hypotonic solutions of various metal chlorides YOSIDA and OGAWA(1956)*, found that the degree of spreading and splitting of metaphase
chromosomes varied according to the sort of cations present in the solutions. The present report deals with the influence of chemical anions on
the chromosomes in the YOSHIDA sarcoma cells.
In order to investigate the effects on anions, 16 kind of Na-, K- and Casalts were used: NaCI, NaH 2P0 4 , Na 2 HP0 4, NaHC0 3 , NazSO a, Na 2S04,
Na2SZ0a and NaF; KCI, KBr, KMnO" K ZCr 207 and K 3[Fe(CN)6]; CaClz, andCa(CH 3C0 2)2' The tumor cells were treated with N/lO and N/lOO solutionsof the above compounds. They were then stained with acetic orcein bymeans of usual squash technique (YOSHIDA and ISHIHARA 1952)**. Among thesolutions of Na-salts, the most satisfactory results for the splitting andspreading of chromosomes were obtained with the N/lOO solution of NaCI
and the N/lO and N/lOO solutions of NaF. For K-salts, the N/lOO solutionsof KCI and KBr showed good results. In Ca-salts the N/lOO solution of
CaClz was the most effective.
Of the compounds studied it can be concluded that the halogen anions
cause the greatest spreading and splitting of chromosomes.
* YOSIDA, T. H. and Y. OGAWA 1956. Ann. Rep. Nat. Inst. Genet. Japan 6: 20.
** YOSIDA, T. H. and T. ISHIHARA 1956. La Kromosomo 29: 1005-1009.

29

RESEARCHES CARRIED OUT IN 1957

C.

DEVELOPMENTAL GENETICS OF ANIMALS

19.

Development of Muscle Protein in Early Chick Embryo(By Yoshito OGAWA, Takatada KAWAHARA and Jiroo MIURA)

As early as on the sixth day of development of a chick embryo, slow
contractions are shown by migrating myoblasis (HERRMANN 1952). This
property of contractility is present in the forming muscle well before theappearance of the usually accepted histological characteristics. In spite of
the primitive morphological stage of early contractile cells, there is evidence that biochemically they have already reached a rather advanced
development (DEHAAN 1956). CSAPO and HERRMANN (1951) found in chickmuscle of the ninth day of development a protein with solubility andviscosimetric characteristics similar to myosin, suggesting the possibility
Table. l.

Numberof
Embryos

Antigen Concentration
Note
4x

24

8x

16 x

32x

64x 128 x~~-----

~

-

_

.

_

.

_

-

-

Hours
after
Incubation

Results of precipitin reaction with actin anti-serumand saline extract of chick embryos.

8
4 Embryos mixed

48

8

72

4

96

+
+

2

*+

2

+
+

-----

144

2 Embryos mixed

±

-Itt

*+

-Itt
-Itt

----

+

+
+

+
+

+
+

-----------

216
Control(Nonfertilized) :2++--------~---

8* + ++* * -Itt*+ * -Itt*+* -Itt*+**--_ .... _ - - - - - - - --------**~.~------

4 Blastodisks
mixed* The heart tissue was carefully excised.
Actin from the heart tissue and that from the skeletal muscle showed no difference in the reaction with anti-serum.

RESEARCHES CARRIED OUT IN 195730

of its existence at still earlier stages. HERRMANN (1952), however, reported
later that muscle protein was not detectable until about the fourteenth day.
We obtained pure actin and myosin from the pectoral muscles ofWhite Leghorns by the method of Szent-Gyorgyi (1944). This protein, at
a concentration of about 15 mg. per mI., was injected intravenously into
rabbits at three day intervals, until it reached a total of 60 mg. The
rabbits were bled by heart puncture on the fifth day after the last injection, and the serum obtained was used for the· precipitin reaction withsaline extracts from chick embryos. A possible failure due to antigenexcess was prevented by using a constant volume of serum and progressively decreasing amounts of antigen.
It was found that actin becomes first detectable in the 72 hours oldchick embryo (Table 1). In view of the fact that heart contractions begin
about 48 hours after incubation, it may be said that the stage of the first
Table 2.

Results of precipitin reaction with mysin anti-serumand saline extracts of chick embyos.

Hours
I Numberafter!
of
Incubation ! Embryos
48

10

72

20

Antigen Concentration
Note
4x

8x

16 x

32 x

64 x 128 x
5 Embryos mixed*10 Embryos mixed
20

96

120

10

+

+

±

*
*+

+

+

-Itt

-Itt

*
*+ *
*
+
*
~o-I--:--:~ **
168

3

!

-tH-

+
+
+

+

I

Control(Nonfertilized) .

*
-*
Itt
*
*
*
*
-Itt

12

* The heart tissue was carefully excised.

+

*

-Itt
-Itt

+
+

* 5 Embryos mixed

-Itt
-Itt
-Itt

*
*

*
* *
* -Itt-Itt
*

*
*

*
*
6 Blastodicks
mixed

RESEARCHES CARRIED OUT IN 195731

contraction of heart muscle corresponds to the stage of actin formation.
(OGAWA, Y. et al.: Kagaku. 27: 253 1957)
On the contrary, in the skeletal muscle, myosin was ascertained about
96 hours after incubation (Table 2).
Myosin in the heart muscle of a chick about 16 hours after incubation
was determined first by EBERT (1953). It may therefore be assumed thatthe appearance of heart myosin, action and myosin in the skeletal
muscle occurs, respectively, about 16, 72 and 96 hours after incubation.
It seems that in the heart myosin appears before the formation of actin,
while the order may be reversed in the skeletal muscle.

20. Promoting Effect of Gonade Extracts on the Spawningof the Pearl Oyster (Pinctada Martensii)(By Yoshito OGAWA, Yushiroo HARADA* and Toshio AI*)

It is a serious problem in rearing pearl oysters that when the gonade isin mature condition, it becomes not only difficult to insert pears kernels
into it, but operating at such a stage spoils the quality of the pearls produced. Though artificial promoting of spawning has been tried by many
workers, no successful method has been reported.
The writers have injected the extract from gonades of Pinctada il1artensii
into the foot tissue of pearl oysters. The extract contained a basic com-!
Contr.
j~eu t ral

Sb.

Acidic Sb.

Basie Sb.

oMay,?

June,5

August ,15June,19
July,7

Fig. 1. Promoting effect of ether-soluble basic compounds
from gonade tissues on the spawning of the pearl oyster.

* Sizuoka Fishers Experiment Station, Simizu.

32

RESEARCHES CARRIED OUT IN 1957

pound which was ether-soluble and non-evaporative. This treatment was
found invariably to bring about spawning about two weeks after injection.
Completly mature and hypertrophous gonades were given grade 5 and thosein faded and reduced condition grade O. The grades between 5 and 0 represented successive intermediate stages (Fig. 1). Thus spawning could
be accelerated by two months and this condition of the injected ovary was
kept for a long time.
This method may help the pearl industry. Chemical purification of theeffective substance is now being studied.

D.
21.

CYTOLOGY OF ANIMALS

Mitotic Activity of Regenerating Liver Cells after
Partial Hepatectomy(By Yoshito OGAWA)

Mitotic activity is often used as a sensitive and reliable criterion tomeasure the degree of growth and regeneration of organs. In our experi·ments with hepatectomy, however, we have frequently met with cases inwhich mitotic activity of regenerating liver cells was not proportionallyrelated to the physiologically regenerating grade. From this point of view,the relation between tissue weight and mitotic activity in the regenerating
liver after partial hepatectomy was investigated using the Wister strain ofrat (No. 1-13, two month old: No. 21-25, 5-6 month old). We measured
the frequency of metaphasic cells 48 hours after operation, observing 2,000
cells per liver lobe.
When more than 60% in weight of the whole liver was taken out by
operation, the remaining tissue greatly increased in weight after 48 hours,
but no mitotic cell was seen. On the other hand, if the part taken out of
the liver was less than 60%, the increase in weight of the remaining partwas relatively small but the mitotic activity was considerably high (Table
1). (OGAWA, Y. et al.: Med. and BioI. 45: 12, 1957)
Further we investigated the relation between the regenerating process
and mitotic activity in each liver lobe, taking less than 60% weight of theliver. As shown in Table 2, the operated lobes decreased but the nonoperated ones increased considerably in weight. However, regarding mitotic
activity no remarkable difference was found between the operated andnon-operated lobes.

RESEARCHES CARRIED OUT IN 1957
Table 1.

33

Regenerating process of liver tissue after partial hepatectomy.

No.

Sex

1

'2)'

2
3

+
'2)'
+
+

4

5
6
7
8
9
10
11
12
13

Treated

+
+

'2)'
'2)'

Body
Weight(g)

W . ht f Increase inRelg~ Weight of
Mitotic
eJ?ove Regenerating
T Issne L'T'•0/
Iver Issue In d ex
III /0
(%)**

-----

-----

90.0
127.8
108.7
129.5
109.3
106.0
96.0
125.6
91.5
129.7
117.1
82.9
113.4

42.8
46.4
35.5
49.6
51.0
50.5
53.6
63.2
63.4
64.6
75.0
75.0
77.3

+
'2)'
+
+
---------84.9
+

----

101

+

102
103
104
105

Control

+
'2)'
'2)'

+ 41.6+ 48.9+ 54.2+ 77.1+ 28.0+145.0+250.9+ 71.8+288.5+ 82.3+162.7+199.0+191.6---------------

1.6
0.0
0.0
1.6
0.0

3.9*
3.5*
3.4*
5.7*
5.0*

85.6
82.5
98.7
105.7

70.0
58.4
46.7
33.4
15.0
8.3
1.6
0.0
1.6
1.6
0.0
0.0
0.0

* Liver weight.
*,' Percent of the whole liver weight estimated.

Table 2.

Variation in the weight of regenerating liver tissue afterpartial hepatectomy.
~~~-

No.

Sex

WeightBody
of
Weight removed(g)Tissuein %II- - -----------------------

21
22
23
24
25

'2)'
'2)'

112.3
121.5
180.5
155.1
175.0

33.6
31.8
34.1
34.1
33.6

+
+
+
-------------c-_~_---

Non-operated Lobes
(%)*

+48.8
+89.2
+46.3
+49.2
+ 1.5

+57.2
+27.1
+41.3
-10.5
+ 2.2

Operated Lobes
(%)*

VI

III+IV

V----_.-----+ 41.5+ 27.3+ 3.1+134.0+ 31.8

-48.7
-41.2
-70.9
-74.6
-56.2

-58.2
-52.0
-58.4
-46.2
-71.4

* Increasing (+) and decreasing (-) percentage were estimated in percent of
the whole liver weight.

34

RESEARCHES CARRIED OUT IN 1957
Table 3.

Mitotic index in each regenerating liver lobs afterpartial hepatectomy.
- - - - - - - - _ . _ - - _ . _ . _ - - - - _ ..

--,----

No.

_---~------_._--_._-----_._.-'-'-'--

Non-operated
Lobes (%)*

Sex

Operated
Lobes (%)*

~ - - ~ . _ -II

VI

III+IV

V

0
2
4
0
1

2
3
1
0
2

2
3
0
2
0

--------

Treated

21
22
23
24
25

0
1
0
2
3

1;1;-

Sf'Sf'Sf'-

Per lob•..

Control

1;-

Sf'Sf'1;1;-

---

7.1

- _ _------

121
122
123
124
125

1
3
2
1
2

0
0
0
0
0

5
12
7
5
8

7.5

~-----'-'---------

1
0
0
0
0

Mitotic
Index

0
0
0
0
0

------'-------

0
0
0
0
0

----

,

0
0
0
0
0

1
0
0
0
0

From these results, we may conclude that:
1) The increase in tissue weight is not always related to mitotic activity in the regenerating liver.
2) Mitotic activity may be influenced by the action of some diffusible
substance which is produced in the regenerating liver tissue.
3) Young rats produce more of such mitotic stimulants than the older
ones. (Table 1 and 2)
For measuring the degree of regeneration of liver tissue by mitotic activity after partial hepatectomy, the extent and place of operation should
be carefully considered.

E.

GENETICS AND CYTOLOGY OF CEREAL CROPS
22.

Ecogenetical Studies in Agropyron(By Sadao SAKAMOTO)( I) A new ecotype of A. tsukushiense var. transiens OHWI
A. tsukushiense var. transiens (2n=42) is a very common plant in Japan,
growing in fields and along road-sides. A strain which differs in several
characteristics from the well known type was found as a swarm in idle
lying paddy fields only, in the hilly vicinity of Misima (Table 1). In comparison with the type, plant height, first internode from top, flag leaf and,35

RESEARCHES CARRIED OUT IN 1957
Comparison of several characters distinguishing A. tsukushiensevar. transiens and the new eeotype.

Table 1.

Experimental field

Natural habitat

Characters
Eeotype transiens

Eeotype

63.0
84.6
117.2
33.4
34.8
53.5
14.4
18.0
17.4
14.8
25.4
26.3
13.3
23.0
23.6
7.6
7.6
7.3
0.80
1.08
0.91
4.29
3.23
3.92
0.84
0.92
1.03
0.49
0.54
0.67
4.8
11.0
2.3
21 May '57 27 April '57
21 May '57 30 April '57

37.9
19.4
7.6
10.2
9.6
5.7
0.65
3.14
0.92
0.60
7.8

transiens--'~.'------------_._-----,.

Plant height (em)First internode from top (em)Flag leaf (em)Spike (em)No. of spikelets
No. of florets per spikelet
Empty glume (em)Lemma with awn (em)Palea (em)Seed length (em)Seed weight (mgm)
Date of first earing
Date of first floweringt._-_._._ ..-spike of this strain are shorter, and the number of spikelets is much smaller but awn, empty glume, lemma, palea and seed are longer, seed weightis much heavier, and the flowering time is about 20 days earlier. Artificial hybrids between A. tsukushiense var. transiens and this strain were
easily obtained and the growth of the F 1 plants was very vigorous.
This strain was recognized as a new ecotype of the above variety of A.

tsukushiense.
Natural hybrids between A. Mayebaranum HONDA and A. tsukushiensevar. transiens OHWI
A. Mayebaranum and A. tsukushiense var. transiens are both hexaploid(2n= 42), but their natural habitats are very different. The former is restricted to moist paddy fields or ditches, while the latter is found alongroad-sides and in fields.
In the sympatrical regions of both species in the suburbs of Misima, 12clones of the natural hybrid between those two were found on the footpaths of the paddy fields. This hybrid found also in Fukuoka Prefecture,
Kyusyu Island, has been referred by taxonomists to A. Mayebaranum var.
intermedium HATUSIMA. Among the 12 clones 9 were non-waxy and very
similar to each other, and the other 3 were waxy. One of the latter wasa dwarf with short spikes. It was considered to be a hybrid with the ecotype mentioned above.
In 1956 artificial hybrids were very easily obtained, and the 17 F 1 plants
were vigorous but completely sterile. The natural and the artificial hybrids
were morphologically similar and resembled A. tsukushiense var. transiens,

(II)

36

RESEARCHES CARRIED OUT IN 1957

but their spikes were not nodding when ripe due to complete sterility.
Chromosome conjugation in meiosis of the natural and the artificial hybrids showed mostly 21n. From this result it is assumed that both species
have the same genomatic constitution. Complete sterility of the hybrid
might be due to cryptic structural differences between the chromosomesof the two species. To throw light upon this point, cytogenetical studies
are now in progress.

23. Morphological and Physiological Variation among
Aegilops squarrosa Strains Collected in Pakistan,
Afghanistan and Iran
(By Hitoshi KIHARA)

176 strains of Ae. squarrosa were collected along a route of over 6000
km in Pakistan, Afghanistan and Iran. The whole area, where Triticum
and Aegilops were collected, is divided into 8 regions from geological aswell as ecological considerations: Quetta, Kabul, Pulikhumri, Maimana,
Teheran, Gorgan, Pahlavi and Tabriz. Morphological studies of thesestrains were undertaken mainly with plants cultivated in the experimental
fields in Kyoto and Misima. Physiological investigations were made witha part of strains representing 8 regions.
Ae. squarrosa is a polymorphic species. Two subspecies, ssp. squarrosa,
and ssp. strangulata, are clearly distinguished. Among ssp. squarrosa, there
are known three varieties: typica, anathera and meyeri. There are many
intermediate types between the first two, which are found almost in thewhole area. Meyeri, which is morphologically an intermediate between
the two, is found solely on the west coast of the Caspian Sea. Ssp. strangulata is localized along a narrow stretch of the south east coast of the
Caspian Sea (Gargan). Its occurrence along the route was estimated tocover 320 km. Ae. squarrosa was found very often in wheat fields.
Therefore it is no wonder that we could find abundant spikelets of Ae.
squarrosa in wheat grains sold as chiken feed. Such characters as erect
culms and large grains, which can be found among cultivated wheats, arefound only among Ae. squarrosa strains collected in wheat fields.
Many morphological variations were discovered. They are related toplant height, growth habit (procumbent or erect), waxy (non-waxy) leaves
and colour of seedlings (red or green) etc. Variations in physiologicalcharacters were also found. They are related to resistance to some ruststrains, earliness and habit, namely winter, intermediate and spring growth.
As to these characters some local varieties seem to have adapted themselves to their habitats. For instance vars. meyeri and strangulata, which

RESEARCHES CARRIED OUT IN 195737

are resistant to certain rust strains, grow in very humid places, along thecoast of the Caspian Sea. All other regions are extremely arid.
Hybrids between strains from 8 regions and also hybrids between threeold strains (Nos. 1-3) on one hand and 8 strains from 8 regions on theother were produced. All F 1 hybrids have shown 7 bivalents. Meiosis isnormal. Fertility is normal in most cases. However some combinations
show lower fertility. Hybrids of strains No.2 give very often rise tohigher sterility. Sterility of the intraspecies crosses seems to depend on
the genotypes of the parents and partly on environmental conditions.
Most probably Iran is the center of diversities of Ae. squarrosa.
The results will be published in detail in collaboration with Mr. M.
TANAKA, Laboratory of Genetics, Kyoto University.

24.

Karyotaxonomic Studies in Poaceae, V(By Tuguo TATEOKA)

1. The systematic placing of two grass genera was studied and theirchromosomes were examined.
Brachyelytrum-This genus includes two species: B. ereetrum Beauv.
which is distributed in North America and B. japonicum Hackel found inJapan and Korea. Several investigators are of the opinion that Brachyelytrum is a member of Stipeae, while other investigators have placed thegenus within Festuceae or Bromeae which are closely related to Festuceae.
B. japonicum whose chromosomes have been examined by the author for
the first time shows 2n=22 in root tip cells, and the chromosomes are
smal!. The chromosome features are very similar to those of Stipeae andclearly different from those of Festuceae or Bromeae. Also the characteristics of leaf structure show a relationship between Stipeae and Brachyelytrum. However, the spikelets of Brachyelytrum have distinct features by which the genus can be easily distinguished from Stipeae. Thus,
it seems natural that Brachyelytrum should be treated as an independent
tribe, Brachyelytreae, which ought to be placed near Stipeae.
Astrebla-This genus comprises four species which are endemic in Australia. According to BENTHAM (1881), GARDNER (1952), a.o., Astrebla may
be referred to Chlorideae, but PILGER (1954) is of the opinion that the genus can be placed within Festuceae together with Triodia, Plectrachne andNotochloa. The present author has observed the somatic chromosomes ofA. lappacea Domin and A. peetinacea F. Muel!. Both species show forty
small chromosomes in root tip cells. Since the basic chromosome numberof 10 is often found in grasses, the chromosomes of Astrebla seem to bea multiple of 10. Not only in the basic number but also in the chromo-

38

RESEARCHES CARRIED OUT IN 1957

some size, Astrebla is very similar to members of Chlorideae and clearly
different from Festuceae. Further, in leaf structure, the Chloridoid subtype of Panicoid type (PRAT 1936) is shared by Astrebla. Thus, Astrebla
should be linked with Chlorideae.
2. After BENTHAM'S monumental treatise (1881), the system of Poaceae
was submitted to studies by many investigators. The present author, also,
has tried to elucidate the natural systematic grouping of grasses. Considering the features of chromosomes, leaf structure, external morphology,
etc., as well as the geographical distribution, the author proposes to divide
the grass family as follows. (ct. Journal of Japanese Botany 32: 275-287,
1957).
Subfam. Pharoideae
Anomochloiformes-Trib. Anomochloeae
Streptochaetiformes-Trib. Streptochaeteae
Bambusiformes-Trib. Bambuseae, Trib. Olyreae, Trib. Streptogyneae
Oryziformes-Trib. Phyllorachieae, Trib. Oryzeae
Parianiformes-Trib. Parianeae
Micrairiformes-Trib. Micraireae
Subfam. Arundoideae
Coleanthiformes-Trib. Coleantheae
Stipiformes-Trib. Brachyelytreae, Trib. Stipeae, Trib. Aristideae.
Glyceriiformes-Trib. Meliceae, Trib. Glycerieae
Nardiformes-Trib. Nardeae, Trib. Lygeeae
Ehrhartiformes-Trib. Ehrharteae
Arundiformes-Trib. Centotheceae, Trib. Phaenospermeae, Trib. Arundineae, Trib. Thysanolaeneae, Trib. Danthonieae
Arundinelliformes-Trib. Garnotieae, Trib. Arundinelleae
Subfam. Pooideae (Festuciformes)-Trib. Festuceae, Trib. Monermeae, Trib.
Triticeae, Trib. Agrosteae
Subfam. Eragrostoideae (Eragrostiformes)-Trib. Spartineae, Trib. Chlorideae, Trib. Lappagineae, Trib. Pappophoreae
Subfam. Panicoideae
Paniciformes-Trib. Lecomtelleae, Trib. Boivinelleae, Trib. Anthephoreae,
Trib. Melinideae, Trib. Trachyeae, Trib. Isachneae, Trib. Paniceae
Andropogoniformes-Trib. Andropogoneae, Trib. Maydeae

RESEARCHES CARRIED OUT IN 1957

25.

39

Fertility Variation due to "Gametic-Development Genes"in Hybrid Populations of Rice
(By Hiko-Ichi OKA)This report is an attempt to explain the change of segregation ratios
and fertility variations observed in back-crosses and F 2 of two distantly
related rice varieties, on the basis of the hypothesis of "Gametic-Development genes". The materials used are strains 414 (P.T.B. 10, Indica type,non-glutinous) and 563 (Kinoshita-mochi, Japonica type, glutinous), whose
F I is semi-sterile.
First, it was found that reciprocal crosses did not differ significantly
from each other in regard to the distribution of fertility in the F 2 andback-crosses (Table 1a).
Secondly, the non-glutinous gene was found to have remarkably increased
against its glutinous allele both in F 2 and back-crosses. From the backcross experiments, the gametic ratio in F I was found to be 80:44 or 1:0.44.
In view of the fact that plants with low fertilities were mostly among
heterozygotes for the glutinous gene, this change in segregation ratio
seemed to be due to the G.D. genes (c.f. OKA 1953, Jap. Jour. Breed. 3: 3139). Assuming that the parental strains 414 and 563 had genotypes + - XI X 2
and gl-x j X2 , respectively, the recombination fraction between + and
Xl or gl and Xl was calculated to be 0.0645.
The frequencies of various
genotypes in the F I gametic series were computed from this value, andthe frequency of semi-sterile plants (due to the G.D. genes under consideration) among hetero- and homo-zygotes for the glutinous gene was estimatedto be 0.483 and 0.061, respectively. Assuming again that the fertility variation due to other causes than the G.D. genes is normal, the fertility distributions in hetero- and homozygous classes were calculated. They were
found to be comparable with the observed distributions, as seen in Table
lb.
Regarding the F 2 , the writer has formerly pointed out (OKA 1953, Jap.
Hour. Breed. 3: 23-30) that pollen with a double dominant combination(X I X2 ) of G.D. genes tends to have a reduced fertilizing capacity (denotedby 1-s). Having computed the value of s from the F 2 ratio of + +: +gl:glgl
plants taking the recombination fraction between gl and Xl (p) to be 0.0645,
it was found that s= 0.836. The observed numbers of + +, + gl and glgl
plants gave a good fitness to the calculated numbers thus obtained fromthese values of p and s. The fertility distribution in F 2 was also calculated by the same method as used in the case of back-crosses, and wasfound to agree with the observed distribution (Table 1c).
These experimental results may serve as an example of successful

40

RESEARCHES CARRIED OUT IN 1957
Table 1.

Distributions of good pollen percentage in hybrids between two
distantly related rice strains, 414 and 563.
-----

% of good pollen
Cross
100

90

80

70

60

50

40

30

No. Meanof s( %) Remarks
20 plant-----a) Comparison betweeen reciprocal crosses:
414 x563 F 2 13
12
10
6
4
3
563x414F 2
6
11
10
4
2
4
(414x563)x563 4(563x414)x563 4

9
3

4
4

12
5

10
11

9
3

2
6

3
2

1
1

54
46

y2=3.08
P>0.98

3

4
1

1

55
32

y2=4.64
P>0.90-------------------b)

Distributions in hetero- and homozygous classes fyr gl gene (in back-crosses
F 1 x563):
Obs.
4
7
6
11
12
9
2
4
5566.4 y2=3.60+gl Exp.
3.37 5.84 9.72 11.93 11.047.573.78 1.73
5563.6 P>0.50
glgl Obs.
Exp.

4
5
2
6
9
3
1
1
1
32 69.1 y2 = 9.38
3.29 5.23 7.50 7.50 5.01 2.440.790.21 0.03 3274.2 P>0.05c)Distributions in F 2 :
7
5
5
+gl
8
14
13
glgl
4
4
2

++

Total
Exp.

2
7
1

2
4

3
4

1
5
2

2
3

2

27 76.2
6072.7
13 82.3

19
23
20
10
6
7
8
5
2
10074.9 y2=13.45
14.47 18.84 17.58 16.63 13.67 9.28 5.44 2.73 1.36 10074.9 P>0.05

explanation of modified segregation ratios and accompanying fertility variations on the basis of the G.D. gene hypothesis.

26.

Back-Cross Experiment between Distantly Related Varieties of Rice-An Interaction between Maternal and Gametic Genotypes(By Hiko-Ichi OKA)

Back-crosses were repeated between two rice varieties, 414 (P.T.B. 10,
Indica type) and 504 (Taichung 65, Japonica type), using the latter as therecurrent parent. The F 1 between these two varieties showed 20% to 24%
fertility in seed setting, and no significant difference was found between
reciprocal crosses. In early generations of the back-cross, fertility ranged
from about 10% to 80%, and plants with 30% to 50% fertility were relatively large in number. In later generations (B 5 -B6 ), most plants showe::l.
a 30% to 50% fertility, and the repetition of back-crosses did not bringabout an improvement in fertility. In about 67% of B5 plants, or 83.5%
of B6 plants, all chromosomes should be of the recurrent parent's origin.
In fact, those plants had almost the same appearance as the recurrent

RESEARCHES CARRIED OUT IN 1957

41

parent 504. Thus, the sterility persisting in those plants seems to suggestthat cytoplasmic effect is involved, as has often been reported in this typeof back-cross experiments with plant species.
Table 1.

Fertility variations observed in a back-cross experimentin rice, (414x504)x504x504···.

% of seed setting
Generation
90-_.-----

80

70

60

~ - - - -Fl
B3 (8 lines)
B5 (5 lines)
B 6 (6 lines)
504 x B5 (6 lines)
B5 F 2 (6 lines)

50

40

30

20

10

8
12

1
6
1

~ ~ - - -- - - ~ . _ -

19

2
18

4
3
11

2
1
3
3
4

~-~---

14
1
14
12

32
16
6
5

3
35
12
10
1
1

No. ofplants
12
101
31
37
26
53

----

However, having used the recurrent parent as the maternal plant in the
back-crosses, the progeny, though it should have the same cytoplasm asthat of the recurrent parent, was still semi-sterile. In contrast, if thosesemi-sterile plants were self-pollinated, the resulting plants were almost
completely fertile without a remarkable segregation in fertflity.
The above-given results can be simply explained by assuming that a pairof genes A:A l are concerned and if A is present in the tissue of the maternal plant, gametes with A l deteriorate. Thus, let the genotypes ofparental varieties, 414 and 504, be AA and AlA l respectively, the F l , AA l ,will have a 50% sterility, and will produce gametes carrying only A. The
gene A:A 1 mayor may not be a group of linked genes. If it were a single
locus, there might be many other multiple allelomorphs which do not bringabout sterility in heterozygotes, and A l might have originated from thoseallelomorphs.
This is a hypothesis alternative to the Gametic-Development gene hypothesis (c.f. OKA 1953 Jap. Jour. Breed. 2: 217-224); the latter is essentially based on 1: 1 segregation of fertile and semi-sterile plants in crossingexperiments of the design (A x B) x C, and change of segregation ratios
accompanied by the predominance of semi-sterile plants among heterozygotes for the given gene pair. These phenomena may however be explained by the new hypothesis. The only phenomenon which the new
hypothesis fails to explain is the restriction on recombination of independent
genes in semi-sterile hybrids, while the G.D. gene hypothesis cannot account for the results of the present experiment.
In addition, the writer has suggested in the last issue of this Annual
Report (for 1956) the presence of "duplicate fertility genes" by which

42

RESEARCHES CARRIED OUT IN 1957

plants with certain recessive combinations have a low fertility. It seems
that we have several different genic mechanisms which form barriers between distantly related rice varieties.

27.

A Preliminary Note on Moist Storage of Dormant Wild Rice Seeds(By Norindo TAKAHASHI and Hiko-Ichi OKA)

It is known in many species of plants that drying of fresh seeds is effective for overcoming their dormancy. In wild rices, however, seeds may
fall on the ground at or before maturity, and may be kept wet in mud
until they get a favorable condition for germination. The seeds may nothave a period of drying for overcoming dormancy, but may need a surviving ability in soil. From this view point, germination of Formosan wild
rice (Oryzasativa sportanea) seeds kept in moist conditions was studied.
1) Moist storage on filter paper in petri-dishes. Fresh seeds of Formosan wild rice lines (mentioned in this Annual Report for 1955) immediately
after harvest, were laid on moist filter paper in petri-dishes, and were kept
at 20°C. Few of them germinated within 30 days. The petri-dishes werethen transferred to 30°C. 38% of those dormant seeds then showed germination within 24 hours. It was found further that, if the seeds were
washed with tap water before transferring to 30°C, the germinating capacity was improved to be 6396. This may suggest that water washing removes some germination inhibitor or metabolite produced in seeds during
the moist storage.
2) Incubation of dormant seeds in soil. Five lines of wild rice whichappeared to have a strong dormancy and four cultivated varieties wereused for this experiment. Fresh seeds of them were buried in loamy soilin pots at the depth of 5 em, and the pots were or were not watered so
as to provide the conditions of ordinary paddy or upland field in them.
They were kept at 30°C or 20°C for 30 days. During this one month
period of soil incubation, wild rices showed on the average only about 5%
germination, while in cultivated varieties about 97% of seeds were germinated. Regarding the temperatures and soil conditions, it was foundthat under the watered paddy condition, 20°C gave higher germination percentages than 30°C, and under the upland field condition, on the contrary,
30°C gave higher germination.
The seeds which had not sprouted in soil were picked up, hulled, andwere tested on moist filter paper at 30°C. They showed quick and complete germination within 24 hours. It may then be said that if dormant
wild rice seeds were buried under ground, their dormancy would be removed, but they do not germinate for some reason and remain alive. ThisRESEARCHES CARRIED OUT IN 1957

43

behavior was particularly apparent when both the temperature and soil
moisture were high. It seems that soil may remove some germination inhibiting substance in dormant seeds in the same manner as water washingof seees in moist storage, but another mechanism may keep them dormant
so as not to exhaust them by immediate germination.
A comparison of germinating capacity of seeds kept under different conditions for 30 days is given as follows:
Incubation in soil (at 20°C)
100 96
Moist storage on filter paper (at 20°C)
38.4%
63.3%
" ,and washing with tap water
Dry storage (at 20°C)
36.4%

F.

CYTOLOGY AND GENETICS OF NICOTIANA ANDSOME OTHER PLANTS
28.

Cytogenetic Studies on the Genus Nicotiana X(By YO TAKENAKA)

The reduction divisions in PMC's were studied in 4 interspecific hybrids:
N. tabacum x N. otophora, N. tabacum x N. longifiora, N. gossei x N. tabacum and N. gossei x N. alata.
1) F 1 of N. tabacum (n=24)xN. otophora (n=12)
At MI in PMC's of F 1 of N. tabacum x N. otophora, the number of bivalents and trivalents per cell was 12 in total. It then seems that the
genome of N. otophora is the same as one of the genomes of N. tabacum.
This finding is generally in agreement with GOODSPEED'S data (1954) on
hybrids between N. otophora and two varieties of N. tabacum.
2) F 1 of N. tabacum (n=24) x N. longijlora (n= 10)
At MI in PMC's of F 1 of N. tabacum xN. longifiora, from 0 to 4 chromosome pairs were found, with the mode at 0-1. The frequency of PMC'swith 2 bivalents followed that of PMC's with 1 bivalent and only univalents. PMC's with 3 bivalents were occasionally found and those with 4bivalents were very rare. Considering such a small number of bivalents
it is difficult to determine whether the few chromosomal affinities are allosyndetic between the genomes of the parents or autosyndetic between
the 2 genomes of N. tabacum. However, KOSTOFF (1943) found usually 5
to 7 bivalents and rarely 8 or 9 bivalents in this hybrid combination. The
cause of the difference between KOSTOFF'S and my own results is unknown.
3) F 1 of N. gossei (n=18)xN. tabacum (n=24)
As far as I know, no report on this hybrid has been published. At MI
of PMC's of this hybrid, the number of bivalents ranged from 0 to 5, with

44

RESEARCHES CARRIED OUT IN 1957

the mode at 1. This pairing is probably due to autosyndetic affinities between the 2 genomes of N. tabacum which is of amphidiploid origin
4) F 1 of N. gossei (n=18)xN. alata (n=9)
The reduction divisions in PMC's of this hybrid were very irregular andappeared to degenerate. At prophase, thin chromatic fibres appeared
around the nucleolus and soon gathered into a clump. Occasionally thin
chromatic fibres spread over the nucleus. Then, in some cells, a large
vacuole appeared in the nucleus which was partly covered by chromatic
membrane. In other cells, a small chromatic sphere or ellipsoid was foundin the nucelus but no vacuole. Only a very small number of cells showedchromosome-like bodies. Pollen grains in the anther were few and varying in shape and size. Moreover numerous powder-like cells and many
non-reduction cells were observed. These observations indicate that N.
alata is not related to N. gossei.

29.

Notes on Seedless Citrus Species(By Kazuo FURUSATO and Yasuo OHTA)

Investigations on seedless Citrus were carried out, and parthenocarpic
fruits were found in species not yet reported as parthenocarpic. They are
as follows:
1) Citrus leiocarpa Tanaka
2) Citrus Hassaku Hort.
3) Citrus madurensis Loureiro
4) Citrus aurea Hort., one strain, Kono No. 436, derived from a seedling of the species.
5) Citrus grandis Osbeck x Poncirus trifoliata RafinesqueCitrus grandis Osbeck x Citrus Unshiu Marcovitch
2 fruits were set following the pollination of 4 flowers in the former
cross and 1 fruit was set from 2 flowers in the latter cross. They
were all seedless.
6) Citrus nobilis Loureiro7) Citrus Natsudaidai Hayata
Parthenocarpy occurred following phytohormone spraying of theflowers.

30.

Trifoliate Seedlings in Citrus(By Kazuo FURUSATO, Yasuo OHTA and Kenji ISHIBASHI)

In general, the three closely related Rutaceae genera, Citrus, Fortunella
and Poncirus, are collectively called Citrus. Among them only one spe-•

45

RESEARCHES CARRIED OUT IN 1957

cies, Poncirus tri/oliata Rafinesque has trifoliate leaves. Wnen pollen of
this species is dusted on the stigma of other species which all have wholeblade leaves, trifoliate seedlings, presumably hybrids, are obtained, but
their leaves show high variation in their shape and size among individual
seedlings.
On the other hand, we obtained many trifoliate seedlings from selfing
or open pollination as well as from pollinations among several species which
do not have trifoliate leaves. They were: C. Tamurana Hort., C. grandis
Osbeck, C. aurea Hort., C. sulcata Hort., C. Hassaku Hort., C. lunos Sieb.,
C. Aurantium L., C. Unshiu Marcovitch, C. Funadoko Hort. and C. sp.
(Kaori-tachibana).
31.

Heteroploid Seedlings in Citrus(By Yasuo OHTA and Kazuo FURUSATO)The occurrence of polyploid plants in Citrus has been reported by several authors. Heteroploid seedling, however, has been reported only one
case. We found many heteroploid seedlings in Citrus which had 1 to 3
extra chromosomes besides the diploid number 18.
Among the seedlings of C. Natsudaidai Hayata, appeared a few (about1%) with slender leaves in addition to some chlorophyll deficiencies as albina and others. Their growth was poor, and they often died in the earlier stages so that in many cases, no root tip could be secured for the examination of chromosomes. The somatic number of 51 plants with slenderleaves were determined, and mixoploid plants were found which had cellswith different chromosome numbers in the same root tip (Table 1). For
the mixoploids the observed 2n numbers are arranged in the table in theorder of their frequency. In Table 1
Table 1. Heteroploid seedlinghetero- and mixoploids are groupedin C. Natsudaidai
together. The table shows that about
Chr. no (2n)No. seedlinghalf of the seedlings were heteroploid,
about 2/3 having 2x + 1 and the remaining18
2~} 2518 19 20
1/3 2x+2 chromosomes. Only one plant19
was found with 2x+3 chromosomes.
19 18
Similar seedlings with slender leaves
19 18 2018
19 20
have been obtained in other Citrus species,
19 20 21
namely C. sulcata, C. Tamurana, C. aurea
19 37
and C. Kinokuni. Heteroploid seedlings
20
20 18
7
seem to be a frequent phenomenon in Citrus.
20 21 36
As to the heteroploids, it is not certain
21
1
1
whether they were of nucellar origin. We _~~_-- -~~__~_~~can only say that two or more heteroploid
Total _ __~~_5~_ _

II}

~}

46

RESEARCHES CARRIED OUT IN 1957

embryos were never found within one seed, but considering their highfrequency, it seems that they may have some relation to polyembryony.

32. Embryoculture in Citrus Natsudaidai(By Yasuo OHTA and Kazuo FURUSATO),

In most polyembryonic Citrus species hybrid embryos are usually small
and morphologicall undifferentiated; they are unable to germinate if sownin the soil by ordinary methods. The present authors have attempted embryoculture with various culture media in order to induce germination of
the so-called small embryos.
Experiment 1: Hydrogen-ion and sugar concentrations
To WHITE'S (1943) solution 2% agar was added. The pH of the solutions
varied from 5 to 7, and sucrose was added in 2, 4 or 8%. The arrangement of 12 experiment batches is shown in Fig. 1. The embryos of C.
Natsudaidai Hayata were separated and classified according to their size
into three groups, i.e. large (about as large as one half of the seed), medium and small (major axis of the embryo shorter than 1 mm). They
were placed in the prepared media and kept in a thermostat at 30°C.
None of the small embryos germinated or became green. Neither did
the majority of the middle size embryos germinate but a part of them became green. All of the large embryos became green and germinated, but
their growth was markedly different, according to the kind of the culture
medium used. The number of leaves, length of stem and main roots after
25 days are shown in Fig. 1.
mm
50

40
30
20
10

I-H--.--r--l--l''-rt"-t-r--rr

10

20
30
40

50

Fig. 1. Growth of Citrus seedlings in culture
media of various pH and various sugar %,the diagram shows the length of stem (above)and root (below), and the flag-like appendages
show the number of leaves.

-60

mm

~02480248!2..J...j..!!.

~

pH

--r-r

5

These results are in accord with the situation in nature. Fruits of C.

Natsudaidai in winter have a pH value of 5 and 11-14 % sugar, but.

RESEARCHES CARRIED OUT IN 1957

47

towards late spring and at harvest time their acidity gradually decreases.
When they ripen and drop off, the content of sugars decreases and theseeds begin to germinate. Accordiogly, for the germinable large embryos a culture medium without sugar but at pH 6 is the most favorable.
Experiments 2: Minor substances
The embryos were separated and classified in two groups, large andsmall. They were placed in the culture media and kept in a thermostat
at 30°C. The culture media contained the following minor elements inaddition to the optimum medium used in expo 1:
Naphtalen acetic acid (NAA)
100, 10, 1, 0.1, 0.01,
ppm
Vitamin B1
10, 1, 0.1, 0.01, 0.001 q
Vitamin C
100, 10, 1, 0.1, 0.01,
q2, 4-Dichlorophenoxyacetic acid (2, 4-D)
10, 1, 0.1, 0.01, 0.001 "2,4, 5-Trichlorophenoxyacetic acid (2,4, 5-T) 10, 1, 0.1, 0.01, 0.001 "None of the small embryos germinated or became green. It was considered, therefore, that the small and morphologically undifferentiatedembryos are also phisiologically underdeveloped and unable to germinate
with the addition only of the minor elements used. The large embryos
became all green. Each of the elements used effected growth repression
in higher concentrations and growth promotion in the optimum concentration. The results are shown in Fig. 2.
mm
40
30
20
10
10

20
30
40
50
60

70
80

~

~

(

I

Fig. 2. Growth of Citrus seedlings in culture media addedwith various kinds and concentrations of minor elements.
The diagram shows the lengthof stem (above) and root
(below), and the flag-likeappendages show the numberof leaves.

In the case of Vitamin B1 the maximum growth was found at a concentration of 0.01 ppm, and the number of leaves in this case exceeded thatof control. This phenomenon agrees with WENT'S report (WENT a.o. 1938)
that Vitamin B1 is necessary for the growth of roots in Citrus and its
optimum concentration is 10- 7 mol.
When 0.01 ppm of Vitamin C was applied, the growth was excellent.
Considering the leaf number, Vitamin C may be one of the promisingelements promoting the germination of embryos and the growth of

48

RESEARCHES CARRIED OUT IN 1957

seedlings in Citrus. This seems to have some relation to high content
of Vitamin C in Citrus fruits.
Thus the so-called small embryos were never germinated. This indicatesthat the small embryos are not only morphologically but also phisiologicallyunderdeveloped and cannot germinate with only the help of the minor
elements used in our experiments.
As to the larger germinable embryos, the growth of the seedlings wasvigorous when the culture media had no sugars and contained Vitamin B1
and C in adequate concentration pH 6.

33. Study on Fiber of tetraploid Luffa cylindrica Roem
(By Kazuo FURUSATO)

4x plants of Luffa cylindrica Roem (var. Turukubi) were obtained bycolchicine treatment. The fiber of 4x was proved to be thicker and heavier
in weight than that of 2x. However, the strength of the fiber was almost
the same in 4x as in 2x in dry condition.

34.

A New Type of Bud Variation in Citrus(By Kazuo FURUSATO)

A bud variation found in Citrus
Unsihu in 1949 was quite different
from the many bud mutations reported up to now for Citrus.
The branch which gave rise to
the observed spontaneous variation
was found on a 40 years old tree.
From its apex, in spring, manylong twigs sprouted forming a nest
similar in appearance to a witches
broom of cherry tree. In winterthe whole structure some time dries
up but in spring the same abnormal
growth is resumed.
The leaves are lighter green andsmaller than on the normal branches.
Flowering was not initiated.
The abnormal branches were grafted on a Navel orange tree asstock.

Photo. 1.
Bud variation in Citrus.

49

RESEARCHES CARRIED OUT IN 1957

The scion retained the abnormal growth and even flowered but the stock
remained unaffected. But when again another precocious Citrus Unsihu
was grafted on the normal scion as stock, some effect upon the former
could be noticed.
The cause of the variation is not yet known. The abnormal growth has
the character of the so-called Inebakanae disease of rice, caused by gibberellin. (Photo. 1.)

35.

Cytological Studies in the Genus Euphorbia. II.
numbers of twenty European species

Chromosome(By Shohachi SHIMOYAMA)The large genus Euphorbia includes about 1,600 species which are distributed in sub-tropical and temperate regions all over the world. They
represent a natural group, although various differentiations can be foundamong them. In 1957, the chromosomes of twenty European species were
examined, and the results are given in Table 1.
Studies on interspecific hybridization are now in progress.
Table 1.

Chromosom numbers of twenty European E7tphorbia species.

Species•nEuphorbia dulcis
E. bivoniae
E. pubescens
E. verrucosa
7
E. gregersenii
E. polychroma
E. polychroma
E. geniculata
E. plathyphyllos
E. exigua
E. exigua
E. virgata
E. heterophylla
E. segetalis . . . . . . . . . . . . .. 8
E. lagascae, 8
E. pilosa9E. amygdaloides
9
E. dendroides
E. Lathyrisl0
E. myrisinites
E. Helioscopia
E. esula.

2nSource12

Hungary
Austria
Hungary
Germany
SwedenHungary
Germany
Hungary
Germany
FranceEnglandGermany
Germany
Germany
EnglandGermany
Austria
Austria
Belgium
Belgium, Germany
Belgium
Germany

24

14
14
14
14
16

28
28
28
56
56
56
16
16
18
18
18
20
20
42

64

50

RESEARCHES CARRIED OUT IN 1957

36. Chromosome Studies in Japanese Callitriche(By Shohachi SHIMOYAMA)The genus Callitriche is a single representative of Callitrichaceae. The
genus includes about 30 species, among which only two are native toJapan. Several species of Callitriche have been studied cytologically by
JORGENSEN (1923), SOKOLOVSKAJA (1932), LOVE and LOVE (1942) and SCHOTSMAN (1954). They have found the basic chromosome numbers of 3 and 5.
The present author has investigated the Japanese Callitriche, and the results which have been obtained up to now are reported in this paper.
(1) Callitriche japonica Engelem
Callitriche japonica Engelem is endemic in Japan. The materials forcytological studies were obtained from different localities: i) Misima, Shizuoka Prefecture, ii) Odawara, Kanagawa Prefecture, ii) Tokyo and iv)
Tomioka and Shimonita, Gumma Prefecture.
The chromosome number of all plants examined is 10 in the root tip
cells without exception, and the somatic chromosomes are of the following
kinds: two large chromosomes with median centromere, six similar medium
sized chromosomes and two small chromosome characterized by a subterminal centromere.
The karyotype can be represented by the following formula.
K(2n)= 10=2Am+6Bm+20t
No meiotic irregularities have been found and all plants examined haveclearly shown five bivalents at first metaphase.
(2) Callitriche verna L.
The materials were collected in rice-fields near Misima City (locatitiesYata, Kawaragaya, Takuchi, Aoki and Shimizu-mura). Their chromosomes
were 2n=20 in root tip cells and 10 II in the pollen mother cells without
exception. JORGENSEN (1923) has counted 2n= 20 in Danish plants belonging to this species, and the same number has been also reported for Russian plants by SOKOLOVSKAJA (1932). Recently, SCHOTSMAN (1954) examined
Dutch materials and observed 2n=20 in root tip cells.
All of the chromosomes of this species have a subterminal centromere,
and they can be divided into three groups according to size; four large(A), fourteen medium-sized (B) and two small chromosomes (C). Two of
the four large chromosomes have each a satellite on the short arm. The
karyotype can be expressed as follows:
K(2n)=20= 2t A l st +2A zst + 14Bst +20 t
Plants collected at Shimizu-mura have no satellite, and their karyotype

RESEARCHES CARRIED OUT IN 1957

51

formula is as follows:
K(2n)=20=4Nt + 14BSt +20 t
In most materials meiotic divisions are entirely normal. But some chomosome irregularities, owing to chromosome stickness have been observed.

G.
37.

BIOCHEMICAL GENETICS OF FLOWERING PLANTS
Leuco-anthocyanin in Variable Flowers of Camellia japomca(By Toru ENDO),

Leuco-anthocyanins are a group of compounds easily converted to anthocyanidin after heating with mineral acids. It is unknown, however,whether those compounds are immediate precursors of anthocyanins invivo. BATE-SMITH (1953) suggested· that leuco-anthocyanin was present
in white flowers of Camellia. The present author found that the flowersof C. japonica contain leuco-anthocyanin which yields cyanidin irrespectiveof the flower color, while the red flowers contain in addition cyanidinglycoside.
A commercial variety of C. japonica, Somekawa, usually has variegatedflowers, but frequently produces red self-colored flowers or branches,
possibly due to the reversion at different stages of a recessive gene effecting white flower color. Out of 176 flowers observed, 66 were selfcolored, two were mosaic and the remaining 108 were variegated. Thisvariety of Camellia seemed to be a good material for studying the actionof genes controlling anthocyanin biosynthesis, since all flowers shouldhave the same genotype except for the mutable gene.
The relative concentration of anthocyanin and leuco-anthocyanin in flowers
having different color types was estimated by spectrophotometrical method.
The red flowers contained about ten times as much anthocyanin as thevariegated ones. Since its spectrum was similar to that of catechin,leuco-anthocyanin was measured together with other catechin-like substance.
The catechin-like substance in red flowers was about 40 % more than thatin variegated ones.
In addition, because of the known importance of copper enzymes inpolyphenol systems, the copper enzyme activity in red and variegatedflowers was estimated by manometrical methods. The results are givenin the following table.
As shown in the table, no significant difference was found between red
and variegated flowers in oxygen-uptake thus estimated. It appears

52

RESEARCHES CARRIED OUT IN 1957
Table
Concentration

Substrate

1.

Enzyme activity
Participating
enzymevariegatedflowers

self-coloredflowers-_._----_._--------

p-Cresol

8 x 10-3M

Catechol2 x 10-3M

p-Phenylenediamine

6 x 1O-3M

Ascorbic acid6 x 10-3M---_._---Polyphenolase
Polyphenolase& Laccase
Laccase
Ascorbase &Laccase

15.7

7.8

20.2

20.9

19.0

15.7

68.0

75.0

----

that copper enzymes, if involved in any way, participate rather indirectly
than directly in anthocyanin formation.

38.

F 2 Segregation of Flower Color in Inter-varietal
Crosses of Swiss Giant Pansy(By Toru ENDO)

Eleven crosses were made among seven varieties of Swiss Giant Pansy,
Mt. Blanc (White), Rhinegold (yellow), Rasberry Rose (reddish purple), Fire
Beacon (yellowish red), Alpenglow (deep red), Lake of Thun (light purplishblue) and Berna (deep purple). One of the crosses (Mt. Blanc x Rhinegold)
was between two acyanic varieties, while the remaining ten were between
acyanic and cyanic ones. The present report deals with their F 2 progeny,
1016 plants in total.
The color pattern of the pansy flower is classified by the relative amountof three pigment systems, i.e. carotenols, red anthocyanins (mainly composed of cyanidin-glycoside), and blue anthocyanins (delphinidin-glycoside).
However, regarding each pigment system, a continuous array of intergrades in color tone was found among the F 2 plants. This seems to indicate that each pigment system is controlled by a polymeric or polygenic
system.
The following results of observations may serve as evidence for theabove assumption:1) In the F 2 of Mt. Blanc x Rhinegold, the range of variation in flower
color seemed to be roughly compared of 33 yellow, 84 pale yellow, and
36 faintly yellowish, though on account of the continuous nature of thevariation the classification could not be a distinct one. In this case, it
was found that none of the F 2 plants had the same amount of carotenol
pigment as the Rhinegold parent.

RESEARCHES CARRIED OUT IN 1957

532) The F 2 of Mt. Blanc x Raspberry Rose appeared to segregate into
42 reddish purple, 15 reddish purple with white stripes, 35 faintly reddish
purple and 35 white with reddish purple parts at the periphery of thepetals.
3) The F 2 of Mt. Blanc x Alpenglow segregated into 20 pale yellow, 56
reddish purple (which might have a smaller amount of carotenol than
Alpenglow) and 16 purplish in which anthocyanins were localized at theouter parts of the petals.
4) The F 2 of Mt. Blanc x Lake of Thun segregated into 69 blue paleblue, 9 blue with white stripes and 3 white which were similar to Mt.
Blanc.
5) The F 2 of Mt. Blanc x Berna segregated into 65 deep purple, 8 bluishpurple and 7 reddish purple.
6) The F 2 of crosses between Rhinegold (acyanic) and five cyanic
varieties showed approximately the same pattern of segregation as thosebetween the cyanic varieties given above, though the number of segregants
with acyanic flower color was larger than in the above crosses.
7) The F 2 of Rhinegold x Berna segregated into 46 deep purple, 59 deep
purple with carotenol in some parts of the petals, and 20 yellowish. In
this case, it seems that the carotenol pigments inhibits the formation ofanthocyanin to some extent.

H.

POPULATION GENETICS AND STUDIES ON
QUANTITATIVE CHARACTERS

39. Genetic Analysis of a White Leghorn Closed Flock Apparently
Plateaued for Egg Production*(By Yukio YAMADA)I.

Time-trends in the means of the population.
The population has been selected for high rate of egg production based
primarily on family average since 1946. The scale of egg production
was transformed to angle from percent production to January 1, as aroutine in analysis of percentage data. The means of the flock were
gradually improved year by year until 1952. Afterwards the means in
1954 and 1956 dropped sharply, even though the means in 1953 and 1955
returned to the same level as the highest mean.
The regression of performance on years was calculated and is given in* This study was carried out at the Department of Poultry Husbandry, Purdue
University, and details will be published in Poultry Sci., 37.

54

RESEARCHES CARRIED OUT IN 1957

Figure 1. The linear regression was significant at 5 % level. The second
degree regression was also significant. This suggests that the rate of
improvement was high in the early generations reaching a maximum in1952 or 1953, after which the means may have decreased. A severe attackof respiratory infection occurred in 1956 resulting in lowered production.
On the contrary, the means of all strains kept at the same farm wereextraordinarily high in 1952. These years could be considered exceptionalfrom the standpoint of environment and may account in part for thesignificant curvilinearity of regression.
Considering all evidence available, it would appear that the curvilinear
trends, as shown in Figure 1, may be exaggerated from the geneticalpoint of view. The linear regression may be considered as more nearly
representative of the time-trends of the means. Nevertheless, the means
in later generations have not increased, indicating that a second degree
regression may actually provide the best fit to the genetic means of the
population. It must then be tentatively concluded that the population hasreached or is approaching a genetic plateau in performance in this population.
70
II. Time-trends in heritability
estimates.
The magnitude of the heritability in a population provides a
Y =52.39+0.89X
measure of additive genetic
variability which determines thea u W M U ~ M Y W
46
47
Year
potential for changing population
Fig. 1. Time-trends in the means of themeans by selection. It has been
population.
postulated that heritability would
decrease during the course of
selection in a closed flock, due to loss of genetic variation by fixation
through selection, and also because of increasing homozygosity from inbreeding, which is inevitable in a population of small size.
The pooled heritability estimated from different components are 0.137
0.239 and 0.188 from sire, dam, and combined components, respectively.
The annual estimates obtained by the variance component technique within
years fluctuate greatly from year to year. So as to evaluate time-trends
in heritabilities, the regression of heritability on year of hatch was calculated. This procedure requires the assumption that there is a linear
relationship between heritability and year, and that the heritabilities
distribute normally at random around the estimated value. Regression
coefficients are listed below, with heritability estimated from the regressions for two years, 1947 and 1956.

RESEARCHES CARRIED OUT IN 1957

SourceRegression
coefficient

h1
h1
h1+lJ

55

Heritability predicted in
1947
1956

t

Probability-.033±.017

1. 93.1O>P>.05

.305

.010

- .024± .011

2.27

.10>P>.05

.373

.153

-.029±.01l

2.65.05>P>.02

.340

.079

Only the regression coefficient for heritability from combined component
is statistically significant. Those for dam and sire components approachsignificance. Although little reliance can be placed on the decline ofheritability from the sire and dam components alone, the fact remainsthat the coefficient of regression for heritability based on the sire component was larger than that from the dam component. This would
suggest a decrease in the additive portion of the genotypic variance, andcould serve as contributory evidence for the hypothesis that the additive
genetic variance will decrease by selection, while, the rest part of the
genotypic variance resulting from non-additive gene effects remains
constant.
Heritability obtained from selection experiments based on averaged
annual selection differential and the annual change in the population
mean was estimated to be 0.08. Compared with those obtained fromvariance components, the value seems quite small. Pooled estimate fromsire component, however, was estimated to be 0.137 which agree fairly
well from the view point of practical breeding, compared with values
obtained by other components.

III. Intended and realized selection intensities.
Selection differential is defined as average superiority of selected birds
over the unselected population from which they were chosen. It wascalculated separately for cockerel, pullet and yearling-hen breeders at
different stages of reproductive cycles, as follows:(h) Intended or initial selection was measured as average superiorityof the mean of birds selected as breeders over the mean for all birds inthe unselected population from which the breeders were chosen. Modification of intended selection due to any association of the performance ofeach selected bird with (1 2 ) the number of eggs set from its mating
during the breeding season, (1 3) the number of chicks hatched from its
mating, (1 4) the number of pullet progeny housed, and (1 5) the number ofpullet progeny completed their records. The last one is the realized
selection. Each selection differential under (1 2)-(1 5 ) was the weighted
means of individual or family superiority over the mean of all birds of

56

RESEARCRES CARRIED OUT IN 1957

the unselected population.
Automatic selection due to association with family size was calculated
as the mean of all progeny minus the corresponding mean of all fullsisters' average. This is the selection automatically applied because thelarger the family size the more the opportunity of selecting the birds tobe breeders.
Automatic selection gave a positive value which means that the larger
families have a greater opportunity for birds to be selected from themthan do smaller families, at least to some extent in this population.
There seems to have a tendency of intensifying family selection for
cockerels and pullets until the stage of progeny housed. Selection for
yearling-hens reached its maximum at the stage of chicks hatched, afterward it reduced to the level of initial selection. As the male breeders
were mated to random samples of selected females, it is reasonable tofind no appreciable changes in the selection differential at the stage ofeggs set. After this stage, however, deliberate selection could be changed,
at least to some extent, because fertility, hatchability and vaibility ofprogeny would be influenced by male breeder's genotypes. Selection atstages of chicks hatched and of pullets housed favored to the male
breeders of superior family record. Realized selection exceeded that ofintended.
Changes in family selection at the stage of eggs set were larger inyearling-hens than in pullets. Fertility or hatchability was not affectedin pullet selection but did in yearlings. Viability favored a little to thebirds which were descendants of superior families in pullets but not inyearlings.
Selection within family in pullets reached its maximum at the stage ofchicks hatched, but selection intensity in yearlings decreased successively
until the last stage of reproductive cycle. The ratio of realized to intended
was 0.75. Superior yearling-hen breeders had poor hatchability and theymight transmit poor genotypes of viability to their offspring.
Selection in males was stronger than that in females and that in yearlinghens was more intense than in pullets. Selection between families wasmuch stronger than that for individual selection within families. The
tendency was very much so in yearling hen breeders.
There was no systematic tendency of the changes in the ratio of realized
selection and intended one during the selection experiment in which the
population had reached a plateau in later half. Intended selection was
realized as intense as it was deliberated.
IV. Effects of inbreeding.
During ten years of breeding the average increase in coefficients of

RESEARCHES CARRIED OUT IN 1957

57

inbreeding among all pullets was approximately 2 percent per year,
although full-sibing was avoided. It has been generally recognized that
rapid or intense inbreeding leads to a decline in fitness. Therefore, itwould be reasonable to expect that inbreeding in this population would
be seriously opposed to the direction of selection.
The technique of regression was used for determining the effect ofinbreeding on performance and size of family. Compulation was carriedout on a within-year basis since this removed the between-year environmental effects on the trait, if there was any.
Of the 10 years considered, only one coefficient of regression of family
average egg production on inbreeding percentage reached statisticalsignificance. The pooled regression was slightly positive but did not approach significance. On the other hand, one regression coefficient offamily size was significant and pooled regression gave negative value
which did not reach any significance level. An additional analysis of theeffect of inbreeding on survivor's production was conducted within-sires
within-year since this would minimize the non-genetic bias. The result
showed that the alternate method has changed the result only slightly
and gave the negative sign for the pooled regression which still was
not statistically significant. Therefore, it seems clear that inbreeding
effect was not seriously opposing the selection applied in this population
if at all.
V. Analysis of effectiveness of selection.
Since selection has been based on superiority of individual, family andprogeny records in this flock, the expected genetic gain per year may be
expressed as follows:,dG= (P- F)h~!n+ (F - P)h;a + (0 - P)h;a'
where P, F, 0 and P are the individual, full-sisters' average, offspring
average and the mean of a population, respectively. The h~!n' h}a and h;a'are the heritabilities corresponding to each selection differential.
For the convenience of calculation, above formula could be changed asfollows:

where ,dS, ,dDp , ,dDp and ,dDo are average realized annual selection differentials for sires and dams.
The results showed that the annual gains realized were not found to besignificantly different from expected gains predicted on the basis of theannual selection differentials and heritabilities estimated from the sirecomponent of variance (P>.20). On the other hand, when the expected

RESEARCHES CARRIED OUT IN 1957

58

values were based on heritabilities from combined sire and dam component,
the differences were significant (P< .05). Thus, the results of selectionin this population cannot be considered to differ from theoretical expectations.
It is reasonable to assume that slight progress could still be made inthis population, since a small amount of genetic variance still remains.
However, such progress would become increasingly difficult to obtain as
indicated by recent generations. Therefore, it is questionable whether
further selection in this population would be worthwhile without someprocedure for increasing the proportion of additive genetic variance.

40. Observation on Natural Populations of Formosan Wild RiceParticularly on Variations in Seed Dormancy
By Norindo TAKAHASHI and Hiko-Ichi aKA)

In this Annual Report for 1955, one of the writers (aKA) has pointed
out that the Formosan wild rice (Oryza sativa spontanea) might be heterogeneous and heterozygous for genes controlling various characters. This
report deals with variations in seed dormancy and other several characters
as a continuation of the former report. Populations A and B, whose
habitats seem to be similar ecologically but are apart from each other by
about one kilometer, were investigated. Plants taken at random fromthe natural populations were grown in 1955 in Misima, and their progeny
lines, 88 in total number, were grown again in this year. All of theplants came to heading in September, but only about one third of them
could reach maturity on account of low temperatures in October in Misima.
Lines from both populations varied in various morphological characters.
The extent of variability within population (C.V.), and the difference between
the two populations as compared with the within population standard
deviation (t), were as follows:-----------------_ .. _- .. _ - - - - - - _ ... ----_ ... -Growing
period(days)

AntherlengthAwn
lengthLigulelength

1.91

0.42
23.0

Tillerangle---------------t*

1.61

C.V.(%)**

10.4

3.16
15.1

LthjWidth
ratio of
grain

49.8

0.33
17.4

3.26
8.1

* t value for the difference between population A and B.
**

Variability within population in terms of the coefficient of variability.

For investigating the variation in seed dormancy, germination wastested immediately after harvest, and the remaining seeds were kept in

RESEARCHES CARRIED OUT IN 1957

59

a desiccater with CaCb at 20°C for further tests. The germination tests
were made under three temperature conditions, i.e., 30°C, 20°C, and dailyalternation between 30° and 20°C. Tests at 30°C generally showed higher
percentages of germination than those at 20°C, and the alternation oftemperature gave the same results as 30°C. However, the degree ofdormancy was shown conveniently by the percentage of germination at
20°C of seeds stored for 20 days after harvest. The results are given in
Table 1.
The table shows that a wide range of variation in seed dormancy exists
in both populations, and population B tends to have stronger dormancy
than A. It is apparent in the table that most of these wild rice lines
have much stronger seed dormancy than usual cultivated rice varieties,
but a few of them seem to come to the extent of cultivated varieties.
Further, correlations between the degree of seed dormancy and othercharacters were investigated. It was found that the degree of dormancy
was significantly correlated with heading date (r= -0.437) and with ligule
length (r= 0.323).
Table 1.

Variations in germination percentage of dorman seedsof Formosan wild rice (within 30 days).

--------

Population.--------

5

15

25

% of germination
35
45
55
65

75

85

95

No. of
lines

4

5

2

3

3
1

2
4

20

------~._.-

A

B

1
1

2
-----

41.

3

1
1

1
2

1

•.. _ - - - - - - - - _ . _ - - _ . , -~-_17---------

Genetical Studies on the Cherry-Red Leaf in Tobacco Plants(By Kan-Ichi SAKAI and Shin-ya IYAMA)In the same manner as in the last year, three lines from each of six
line-groups were raised for the observation of cherry-redness in leaves.
The experiment was designed according to the randomized block methodwith two replications, each plot containing 12 plants (First experiment).
In addition, an experiment with F r and F 2 -hybrids between lines withhigh and low cherry-redness (2 cross-combinations) was made with three
replications, each parental, F l - and F 2 -plot containing 30, 15 and 45 plants,
respectively (Second experiment). Five top leaves were sampled fromeach plant, and after drying, were classified into five classes, 0 to 4, by
visual judgement. The index-numbers 0 to 4 were then weighted by theweighting formula which was calculated from the data of the first experiment, to maximize the ratio of between-plant-variance to within-plant-

60

RESEARCHES CARRIED OUT IN 1957

variance.

The formula obtained for this year was
X=Oxo+0.256xl +0.451xz+0.809x3+ 1.000X4 ,where Xl represents the number of leaves falling on the i-th class ofcherry-redness.
Analysis of variance of the score X in the first experiment showed thatthe variation among line-groups was significant but that within group was
insignificant (Table 1). The heritability value for line means computedfrom variance components was 0.699. With the data for the last three
years (1955-57), analysis of variance was made again (Table 2).
Table 1.

Analysis of variance of cherry-redness in 18 lines ofBright Yellow tobacco (1957).

Source of variationd.£.

Mean square

Replication
Line-group
Line within line-group
Error

1
5
12
17

0.050,161
0.142,177**
0.014.630
0.017.853** Significant at 1 % level.
Table 2.

Analysis of variance of cherry-redness with the pooleddata for 1955. 1956 and 1957.

Source of variation
Line
Year
Year x line
Error
**

d.f.

Mean square

17
2
34
72

0.200.218**
0.101.857**
0.015.867
0.016.575

Significant at 1 % level.

Table 2 shows that the variance among lines was significant as expectedfrom the results obtained in each of the three years, but the interaction
between year and line was insignificant. Further, the correlation coefficient between line means in 1956 and in 1957 was as high as 0.876, andwas highly significant. It may then be safely concluded that the cherryredness is primarily genetically controlled.
The results of the second experiment are given in Table 3.
Table 3 shows that the occurrence of cherry-red leaf in F 1 is at thesame level as that in the parent with higher occurrence, and the F z mean

61

RESEARCHES CARRIED OUT IN 1957
Table 3.

Comparison between F 1 and F 2 of reciprocal crosses
regarding cherry-redness. (1957).

Line

Cherryredness

(1 )

0.321

(7)
F 1 (l)x(7)F 1 (7)x(l).598

F 2 (l)x(7)F 2 (7)x(l)----_..

~~-_._--

Average ofreciprocals.606 }.615.611.624 ).639.654 fLine

Cherryredness

(1)

0.321

(15)
F 1 (1)x(15)F 1 (15)x(l)F 2 ( 1 ) x (15)F 2 (15) x ( 1 )Average ofreciprocals.595.622 }.541.582.574 }
.500

.537

---,.

is near to the F 1 mean. Thus, cherry-redness appears to be dominant inthese crosses.
Correlation between cherry-redness and the quality of tobacco leaf wasinvestigated using the data of the first experiment. The quality of leaves
was evaluated irrespective of cherry-redness for this purpose. The
phenotypic correlation was 0.404 (significant at 1 % level), and the genetic
and environmental correlation coefficients were computed to be 0.085 and0.326, respectively. In view of the low genetic correlation coefficient,cherry-redness may be regarded as a character independent of other factors
relating to quality in selection. The relatively high values in phenotypicand environmental correlations might be due to the tendency to understimating the quality of a leaf if it showed cherry-redness.

42. Genetic Correlations of Agronomic Characters in Rice(By Shin-ya IYAMA)

Genetic and environmental correlations between various agronomic
characters were estimated from the analysis of components of variancesand covariances. The following two groups of materials were used.
Group 1. 19 varieties cultivated in Japan. The experiment was designed
according to the randomized block method with six replications. Plant
height, panicle length, tiller number, panicle number, plant weight, panicle
weight per plant and 100 grain weight were investigated. The genetic
and environmental components of variances and covariances were estimatedfrom the data, and correlation coefficients were calculated as given in
Table 1. The correlations between these characters and grain yield inthree-plant-per-hill cultivation are also shown in Table 1.
As shown in Table 1, tiller or panicle number showed a positive genetic
correlation with yield, but panicle length showed a negative one. This
may imply that among the Japanese varieties used, the so-called "panicle-

62

RESEARCHES CARRIED OUT IN 1957

number types" are high yielders. However, varieties with many panicle
tend to have a small grain weight, judging from the genetic correlation.
In addition, positive genetic correlations were found between panicle
length and plant height, and between panicle number and tiller number.
Genetic correlation between panicle length (or plant height) and tiller (orpanicle) number was negative, but the environmental correlation was
positive, resulting in non-significant phenotypic correlations.
Grain yield of the varieties in three-plant-per-hill cultivation showed asignificant positive correlation with panicle weight per plant. This meansthat selection from single planting should result in obtaining strains which
will have a high yielding capacity in the usual planting conditions.
Table 1. Genetic and evironmental correlation coefficients in rice.
(Genetic correlation coefficients are above the diagonal and environmental
ones below the diagonal. Correlation coefficients from Group 1 are givenin the upper lines and those from Group 2 in the lower lines.)
Plant Panicle Tiller Panicle Plant Panicl.e
10?
height length number number weight ~~.j~~~~~ ;:~~t Yieldt
0.594 -0.334 -0.363
.595
.088
.080

Plant
heightPaniclelength.159.337

Tillernumber.347 -0.043
.555
.281

Paniclenumber.594 -0.045
.566
.212

.458
Plant
.674
weightPanicle.493
or grain.448
wt.jplant
100 grain -0.076
weight

.179
.544

.034
.235

-0.420 -0.423 -0.206 -0.309-0.147 -0.164
.285
.201
1.000
.996

.94~ ,.042 -0.066
.088
.317

.301
.252*

.657
.448

.529 -0.325
.090

.493*

.583
.409

.513 -0.318 ,
.083

.487*

.961 -0.057.643.743**

"

.177
.122

.947

",

.124
.415

.707
.837

.712
.869

.151
.257

.561
.524

.626
.582

'8~,,~:147.833**.699.013 -0.024 -0.020

.144

.215

.~

Grain+
shape.264**
.168

-

.148

+ Phenotype correlation based on line means.
t Yield per plant of three-plant-per-hill cultivation.
* Significant at 5 % level.
** Significant at 1 % level.

Group 2. F 7 lines from Omachi x TeTep. 112 lines were taken at random
from the Fa bulk population of the cross. They were tested in an experiment arranged according to the randomized block design with three

63

RESEARCHES CARRIED OUT IN 1957

replications. Genotypic and environmental correlations among various
agronomic characters were estimated by the same method as used forGroup 1. The results are also given in Table 1.
Table 1 shows that there are essentially the same tendencies as foundin Group 1. But, there are some important differences in that plantheight or panicle length shows a positive genetic correlation with grain
yield per plant, and does not show a negative genetic correlation withpanicle or tiller number. This seems to suggest that in Japan, such typesof rice as showing dwarf habit and high tillering ability have been selected,
resulting in the absence of correlation between panicle length and yield,
and a negative correlation between tiller number and panicle length.
These features may however be limited only to Japanese rice varieties.
Considering the whole pattern of variation among rice varieties, bothtiller number and panicle size are important as yield components.
Finally, regarding the shape of grain, Omachi has apparently shorter
grain than TeTep (length/width ratio were 2.17 and 2.95 for Omachi and
TeTep respectively). A wide variation in grain shape was found among
the F 7 lines. The correlations found between grain shape and othercharacters indicate that lines with a long grain tend to have a long panicle
and heavy plant weight. But grain shape showed no correlation with yield.
Heritability values: Based on the values of genetic and environmental
variances obtained from the above experiments, heritability values forvarious characters were computed as given in Table 2. There were nomarked differences in heritability values between Groups 1 and 2.
Table 2.
Heritability
forPlant Panicle Tiller Panicle Plant Panicl.e
100
height length number number weight °tr /grlam grain Yield*
w . p an t weight

Plant
Plot mean

.460
.809

.348
.748

.393
.851

.372
.840

.280
.740

Plant
Plot mean

.363
.899

.247

.245
.771

.158
.746

.137
.713

Group

2

Heritability values of the characters investigated.

.837

.172
.646

.500
.775

.079
.523

.061
.362

* Grain yield per plant of three-plant-per-hill cultivation.

43.

Migrating Activity in Inbred Lines derived from Two Wild
Populations of Drosophila melanogaster(By Kan-Ichi SAKAI, Takashi NARISE, Toshitaka ITO and Shin-ya IYAMA)

Two wild populations of Drosophila melanogaster, viz. KN and TB popu-

RESEARCHES CARRIED OUT IN 1957

64

lations (for details see the last issue of this report), which differedmarkedly from each other with respect to migrating activity were usedin this study. Fifteen inbred lines have been derived from each of thetwo populations. They were investigated as to their migrating activity
in every inbreeding generation. The method of investigation consisted incounting the number of flies migrated from the original tube 24 hours
after the establishment of an experimental set of population-tubes. The
Table 1. Migrating activity of 12 inbred lines derived from two wild populationsof Drosophila melawgaster in sixteen consecutive generations of inbreeding.
Migrating activity in 16 inbred generations

Inbred
lines

1,2

3,4

5,6

7,8

9,10

11 ,12

13,14

15,16 Average

A-2
B-1
C-3
KN D-2
D-3
F -2
H-l

50.63
53.13
42.71
44.79
71.46
27.92
22.88

45.63
41.46
33.34
49.59
60.21
58.75
45.21

28.05
60.21
25.83
55.04
80.80
37.09
18.34

37.92
77.71
58.96
72.09
49.38
55.63
43.00

40.63
48.34
44.79
30.84
45.21
35.21
23.96

37.71
63.96
56.88
36.25
48.75
27.71
25.83

49.38
64.58
57.71
27.09
52.50
23.34
25.84

61.88
64.58
65.00
31.46
39.59
39.38
16.25

A-I 50.42
B-3 18.75
TB C-2 24.59
C-3 27.50
F-l 27.83

26.46
20.84
48.54
49.58
17.08

28.33
50.63
49.58
49.79
37.09

16.88
50.84
64.38
66.63
39.79

25.00
36.46
43.54
43.96
21.88

22.09
42.30
56.46
66.25
18.34

18.54
46.25
41.88
57.29
14.59

25.83
41.67
51.04
77.50
22.29

26.70
38.47
47.50
54.81
24.86

Average

41.39

43.40

52.77

36.65

41.88

39.92

44.71

42.41

-~

38.55

--- - - - - - -------

Table 2. Analysis of variance of migrating activity of twelve
inbred lines derived from two wild populations for the last
seven inbred generations from 10th through 16th.
-------

Source of variationd.f.

Mean square

Generation
Line
Betw. population
Within population
Generation x line
Error

6
11
1
10
66
168

0.236,157**.534,690**.042,287.583,931**.026,750.027,646* Significant at 1 % level.

43.98
59.25
48.15
43.39
55.99
38.13
27.66

RESEARCHES CARRIED OUT IN 1957

65

number of flies tested in each plot was 80, replicated three times. The
inbreeding was continued up to the 16th generation, during which thenumber of inbred lines were reduced to 7 in KN population and 5 in TB
population. The migrating activity of 12 lines from the two populations,expressed in percent of migrated flies to the total number is presented inTable 1.
Analysis of variance was applied for the data obtained for the last
seven generations (Table 2).
From the two tables given above, we find the following facts: (1) The
migrating activity of flies did not show any definite decreasing tendency
due to inbreeding, though there was a statistically significant variationfrom generation to generation. (2) The difference between two populations
was not statistically significant, but intra-population variation was highly
significant. (3) Interaction between generations and lines was insignificant.
These facts indicate that the same population involves genotypes responsible either for high or for low migrating activity which will be separated
by inbreeding. Despite our initial expectation, the inbreeding has notbrought about a marked decrease in migrating activity.

44.

Variation in Competitive Ability among F g Lines of the Cross
between an Upland Rice Variety and" Red Rice"(By Shin-ya IYAMA)

To inquire into the genetic behavior of the difference in cor ']JetltlYeability between the upland rice and" Red rice ", F g lines of Norin-Mochi
18 x" Red rice" were tested in a competition experiment desif ned asfollows: One of the parents, Norin-Mochi 18, was used as the tester forcompetitive ability. Each of 23 F g lines were mix-planted with be tester
variety alternately in a row. Various characters of the tester plants
were measured, and the difference in row means between mix- and pure
plantings was used for measuring competitive ability of the Fa lines. The
rows were arranged according to randomized block design with four
replications. Plant weight, plant height, panicle length, tiller number,panicle number and panicle weight per plant were investigated. Variations
in row means in mix-planted plots, as compared with the means in purestand, are shown in Table 1.
The results of variance analysis of the data are given in Table 2.
Table 2 shows that the variations among lines in plant weight, panicle
number and panicle weight per plant are significant. It is apparent from
Table 1 that though "Red rice" is the strongest competitor, most Fa

66

RESEARCHES CARRIED OUT IN 1957
Table 1.

Variations in characters of the tester variety mix-plantedwith different Fa lines.
Plant weight

13

15

17

19

21

1

6

7

5

3

23gr.

No. of lines
23

N**

R*
Panicle number
4.0

5.0

4.5

5.5

6.0

6.5

7.0

7.5

8.0

No. of line-------~~~-~-----

8

4

5

2

1

2

23

N

R

Panicle weight per plant

4

5

6

7

3

8

7

3

8

9

10

R:

No. of lines

1

23
N

R

*

llgr.

Mixed with" Red rice".

Table 2.

**N:

Pure stand of Norin-Mochi 18.

Analysis of variance of competitive ability (difference between
mix- and singly planted plots) for Fa lines.
Mean square

Source ofvariationd.£.

Replication
Line+

23

Error

69

*

3

I

Plant
heightPaniclelengthTillernumber

32.2353

14.6658

20.9282

1.599216.0051

1.1746Significant at 5 % level.
+ Including" Red rice".

**

PaniclenumberPlantweightPanicle wt.
per plant

6.7516 3.1215*
2.4536 2.4570*

120.6994**
28.0128*32.6469**

1.781615.6635

2.26661.33036.5376*Significant at 1 % level.

lines are nearly as strong as "Red rice". It may then be inferred thatthe progeny of crosses between upland rice and "Red rice" tends tohave a strong competitive ability and to grow in numbers if mixed witha population of upland rice varieties.

67

RESEARCHES CARRIED OUT IN 1957

45. Genetic Variability in Wild Populations of Oryza
perennis and O. rufipogon(By Kan-Ichi SAKAI and Takashi NARISE)

Oryza perennis and O. rufipogon (or O. sativa spontanea) are considered
to be the very close relatives of cultivated rice species, O. sativa, andare distributed sporadically in the Ceylon island. The former species ismostly found in the western part of the island, while the latter in theeastern part. Both species grow in swamps, on the fringe of water-courses,
and in paddy fields.
A number of panicles were collected from three populations of eachspecies, and ten spikelets were taken at random from each panicle.
Measurement was made on awn length and on the length and width of
spikelet. The places where O. perennis was collected were Veyangoda,
Yagoda and Bombuwela, and the places where O. rufipogon was collected
were I1luppaiya-dichchenai, Pottuvil and a spot near the latter village.
Materials were always collected from a fairly large population.
Mean values for the length and width of spikelets and awn length on
an individual panicle basis are presented in Table l.
The data thus obtained were analyzed by the method of analysis ofvariance in order to estimate the genetic and environmental components
Table 1.

SpeciesO. perennis

O. rujLpogon

Mean values of length and width of spikelets and awn lengthin wild populations of two Oryza species.
Awn
length (em)Population No. of panicles
investigated

Spikelet
Spikelet
length (mm) length (mm)Veyangoda

52
55
67

8.15±0.340 2.60±0.155 6.82±0.800

8050

7. 96±0 .230 2.61 ±0.099 9.31 ± 1.133
7.94±0.253 2.67 ±0.115 7.30±0.973
8.62±0.311 2.75±0.09410.93±1.661

YagodaBombuwela
PottuvilQ(nearby)IIluppaiya.

67

8.25±0.297 2. 76±0.106 8 .59± 1. 771
7.95±0.282 2.44±0.048 9.94±0.843

of variances and covariances. Coefficients of genetic and environmental
correlations were also calculated. In this analysis, it was assumed thatvariation within panicles (among spikelets on the same panicle) represents
the environmental variation, and that among panicles is composed of theenvironmental variation of the same magnitude and the remaining genetic
portion (each panicle represents an individual plant). This assumption is

68

RESEARCHES CARRIED OUT IN 1957

acceptable for the length and width of spikelet empirically. In awn
length, though the environmental variance among plants may not always
be comparable with that within plants, the assumption may still go so as
to yield a preliminary insight.
Genetic and environmental correlations of spikelet length with spikelet
width, and with awn length, are given in Table 2.
It is seen in Table 2 that length and width of spikelet are not correTable 2. Environmental and genetic correlation coefficients between spikelet
length and other characters in wild populations of two Oryza species.
Spikelet length andSpikelet width
Awn length
re
r"
re
fa

SpeciesPopulation

O. perennis

YagodaBombuwela

0.16230.12940.3058-0.0076
0.16190.00570.49530.1926
0.92390.48160.03250.3866

Pottuvil"(nearby)
Illuppaiya.

0.02090.20630.46690.34910.03890.18980.31460.1381
0.39300.48380.65390.7012

Environmental correlation.

r y:Veyangoda

O. rujlpogonre:Table 3.

Genetic correlation.

Genetic and environmental variance components and heritability
values (h 2 ) in wild populations of two Oryza species.
--------.-

O. perennis
VayanBombuYagoda
goda
wela

O. rufipogon
Illupnear
Pottuvil Pottuvil
paiya.

-------------

O'e

Spikelet
length

2

O'y2

h2

0.05900.082758.36

0.10440.026620.31

0.05680.040041.32

0.02680.050565.33

0.04120.027239.77

0.10270.086645.75

0.01560.019956.06

0.00430.002940.28

0.00400.009470.15

0.01200.007137.17

0.00440.050491.97.. _ - - _ .... _ - - - - - _ .. _ - O'e

Spikelet
width

2

0' y2

h2
----------------------.-----------

rYe?.

Awn
length

0.00360.0141
79.66

O'y

h2

2

0.76401.382964.41------% of genetic
variance inaverage-

0.96290.512434.73

1.13530.582933.92

0.11901.030489.65

0.35400.911072.02

0.50871.8027
77.99

37.03

38.51

75.04

49.65

71.90

-~

67.48

----------------_.-,-------RESEARCHES CARRIED OUT IN 195769

lated genetically, or, if correlated, only slightly. In contrast, spikelet
length and awn length are correlated significantly. Of special interest in
this connection may be the very low genetic correlation between spikelet
length and awn length in the Yagoda population, unlike the other five
populations.
The values of genetic and environmental components of variance, andthe heritability value (genetic variance in per cent of the total variance)
were estimated as given in Table 3.
Looking at Table 3, we find that among three populations of O. perennis,
the Yagoda population is genetically the most heterogeneous, and in O.
ruJipogon, the Pottuvil and Illuppaiyadichchenai populations appear to beheterogeneous to the same extent.
From the results of investigations described above, the following conclusions will be drawn: (1) The two species cannot be distinguished fromeach other in so far as spikelet measurements or awn length are concerned.
(2) There is a large inter-population variation within the same species.
(3) Though length and width of spikelets are not correlated genetically,
spikelet length appears to be genetically correlated with awn length. (4)The magnitude of genetic variability seems to differ from population to
population.

46.

Heritability of Grain Shedding and Other Characters in Rice
(By Kan-Ichi SAKAI and J. J. NILES*)This report deals with the estimation of the heritability values of afew of economically important characters of rice in Ceylon. The method
adopted is the estimation of genetic and non-genetic components of variation by the method of analysis of variance in Fa lines. The materials
used were hybrid populations between Pachchai perumal or Panduruwi
and Vellai Illankalayan.
The method of analysis is as follows: Let the number of Fa lines be
x, the number of individuals within each line be y, the number of individuals within a parental (pureline) plot be z, and the number of thosepureline varieties be r. On the basis of measurements taken on each individual Fa plant, the analysis of variance table can be constructed asfollows:* Department of Agriculture, Peradeniya, Ceylon.

RESEARCHES CARRIED OUT IN 195770

Source of variation
Between lines
Within lines
Within purelinevarieties

Degree offreedomMeansquarecomponents of meansquare expected

x-I
x(y-I)r(z -I)~ue,

In this table, ae' stands for environmental variance, a' includes besides
environmental variance, genetic variance due to segregation of genes
within Fa lines, and as" is the genetic variance due to genetic differences
between lines. The genetic variances involved in a 2 and a s2 are expected,
for Fa hybrid lines of autogamous plants, to be.

andwhere a g ' and an' are the variances due to fixable and non-fixable effects
of genes, respectively.
The heritability value of a given character in the Fa generation can be
estimated by

The heritability value of the same character in F 2 can be estimated by

Heritability values were computed by using these formulas. The regression of Fa progeny lines on the F 2 parents, and the number of effective
factors (by Panse's method), were computed further. The results thus
obtained for degree of grain shedding, * panicle length and plant heightare given in Table 1.

RESEARCHES CARRIED OUT IN 1957

71

Heritability values, F 2 -Fa regression, and number of effective
factors for grain shedding and other characters.

Table 1.

Fa

F2Regression of
Fa lines on
F2 parents

PIXV
P 2 xV

0.093
0.392

0.053

0.082

6

Panicle length

PIXV

0.069

0.300
0.041

0.239
0.155

11

Plant height

PIXV

0.612

0.481

0.465

3

PI, P 2 , and V:

See text.

Character

Cross

Grain shedding"

Heritability value

No. ofeffective
factors2* The degree of grain shedding was assessed by running a glass roller overeach of the panicles to be tested on an inclined board. The data given inpercentage were transformed into an index-number before analysis.

1.

I.

RADIATION GENETICS IN ANIMALS

47. Radiation Mutagenesis in the Silkworm
Changes in Sensitivity of Silkworm Germcells to X-rays at
Different Stages of Gametogenesis(By Yataro TAZIMA and Kimiharu ONIMARU)

Marked differences between different developmental stages have been
revealed in the sensitivity and mutability of the spermatogenic cells of
Drosophila and mouse treated by X-rays. In both animals germcells in
different phases exist together in the same testis and the response pattern
was determined by testing successive broods at a definite interval after
irradiation. It may, however, be difficult to tell exactly which type of
spermatogenic cells were tested when the adult males had been irradiated.
The investigation can be more exactly carried out in silkworm which
offers the advantage of almost simultaneous development of the germcells
in the gonads in accordance with the development of the individuals, bothmale and female. Therefore, the present work was undertaken with thesilkworm.
Wild type females and/or males were irradiated with 250, 500, 1000,
2000, 3000 and 4000 r at several stages of larva and pupa and were matedto non-irradiated double recessive pe re partners. Five days after being
laid, the eggs were covered with pigments according to their genotypes.
They were counted seven weeks later, so as to make it easy to distinguish

RESEARCHES CARRIED OUT IN 195772

pe mutants from unfertilized eggs and early embryonic lethals.

The
results were as follows.
1) Changes in the number of eggs laid per mother
By the irradiation of the female with 1000 r X-rays, no noticeablechange was observed among groups irradiated at various stages, but whenirradiation dose was higher than 2000 r a noticeable decrease was foundin the egg number among groups irradiated early. Irradiation at latepupal stage showed no appreciable decrease in the egg number, even at
3000 r.
Male germcells, however, responded quite differently to X-rays fromthose of the females. A marked sensitive period was revealed in theearly fifth stadium when the germcells were in early spermatocyte stage.

~300~- - IODOr_

IV2 lV4

VI

V4

V6

Sl4gesV8

qf males

PE

PM

2000r

PL

irrad iatedFig. 1. Average number of eggs laid per female matedwith males irradiated at different developmental stages.

In this stage decrease in the number of eggs was clearly observed evenat 500 r.
2) Frequency of unfertilized eggs and early lethals
No affected stages were found when females were irradiated with 1000 r,
while in the male a remarkable difference was observed with the same
dose of X-rays according to the irradiated stage. The incidence of unfertilized eggs showed its peak at early fifth stadium, which coincides
with the most sensitive stage in respect to the egg number. The
spermatogonium in the third larval stadium is fairly resistant to X-rays
as well as the mature sperm in the pupal stage.
Thus the sensitivity of germcells to X-rays was revealed clearly inboth sexes of the silkworm. It is noteworthy that there is an extraordinary sensitive stage in early fifth stadium of the male when the
germcells are very badly affected.

73

RESEARCHES CARRIED OUT IN 1957
%
100

l'"
~

...'"

50

~

:&~

-...

§~

.....

"

~

9
O/V2 IV4VI

V4StagesV6

VBof mal",N4 PE
PMand [emaws ,rraaiated.

PL

Fig. 2. Irradiated stages of males and females and ratesof unfertilized eggs.

II.

48. Radiation Mutagenesis in the Silkworm
Mutation Response Pattern of Silkworm Germcells to
X-rays According to Stage and Sex(By Yataro TAZIMA and Kimiharu ONIMARU)

Since conspicuous differences were found in the sensitivity to X-rays ofsilkworm germcells at different stages of the gametogenesis, a similar
detailed analysis of mutation response was made in respect to the rate ofrecessive visible mutations at specific gene loci and that of dominantlethal mutations at various developmental stages of the gametogenesis.
1) Recessive mutation rate at specific gene loci
Experiments were carried out by the same method as mentioned in
the foregoing article. According to this method most of the F 1 eggs are
expected to be of wild type, but some eggs may show recessive mutant
characters of the non-irradiated parents as a result of recessive point
mutation or deficiency at marked loci. From the number of exceptional
eggs mutation rates were calculated at each irradiated stage and plottedwith the developmental stages of the parents.
The mutation rates in the male germcells showed remarkable differences
according to the developmental stages of the irradiated individual, while
the female germcells showed mostly the same results at different stages
prior to mid-pupal stage but after this stage sensitivity suddenly increased.
2) Dominant lethals
The incidence of dominant lethals was analysed by the percentage of74

RESEARCHES CARRIED OUT IN 1957

~V2 IV4VI

V4 VB VI!
N4 PE
PM
Stages of males and females irradiatedPI.

Fig. 1. Change in mutability with the development of
Silkworm germ-cells.

dying embryos per total number of fertilized eggs at four different developmental stages, i. e. stadium 4 day 2, stadium 5 day 2, stadium 5
day 6 and late pupal stage. The response pattern of dominant lethals
was mostly the same as that of recessive visible mutations at specific loci,except that higher doses than 2000 r were required to reveal the differences
in sensitivity. Irradiation group at stadium 5 day 6 showed higher mutation rate than at stadium 5 day 2, which was consistent with the mostsensitive stage with regard to the visible recessive mutation but inconsistent with the stage of lowest fertility. To discuss this point a morethorough investigation is required.
If these results are referred to the stages of gametogenesis, it may beconcluded that the germcells of the silkworm are most sensitive to X-rays
in some stages of spermiogenesis and oocyte, while oogonium, spermatogonium and mature sperm are less sensitive.

III.

49. Radiation Mutagenesis in the Silkworm
Comparison of Xray Induced Mutation Rates in the Silkworm
with Those of Drosophila and Mouse(By Yataro TAZIMA)

X-ray induced recessive mutation rates were calculated from the dataobtained from mutations at loci +pe and +re, and compared with thoseof Drosophila and mouse as follows.
It is evident that visible mutation rates in the silkworm are remarkably
higher than those found in Drosophila. The former are about 7 times ashigh in mature sperm and 28 times as high in spermatogonium as thelatter. It is, however, necessary to bear in mind that in the silkworm

75

RESEARCHES CARRIED OUT IN 1957

Mutation rates per r per locus
Spermatogonium

Bombyx
DrosophilamelanogasterMouse

4.2xlO- 7
1.5 X 10-8 (Alexander '54)

Mature sperm
4.5 X 10- 7
6.0x10- 8 (Alexander '54)

2.5x10- 7 (Russell '51)

mutation rates were calculated with respect to mutant characters expressedin the egg stage in contrast to those observed in Drosophila which wereobtained with respect to adult characters. Actual incidence of mutation
in Drosophila might be much higher than the above data if earlier
characters were tested.

50.

Tissue Culture Analysis of Delayed Lethal Irradiation
Effect in D. melanogaster(By Tsutomu SUGAHARA and Masakatsu HORIKAWA)

It is well known that the effect of radiation on Drosophila larvae is
not their immediate death but a delay of pupation and a decrease of
pupation- and imagination rate in a wide range of radiation doses. Furthermore, it was observed by the authors that the body weight of irradiatedlarvae, whose pupation was delayed, did not only not decrease but wasconsiderably larger than in the control larvae. In the present investigation,
the mechanism of this phenomenon has been studied in some detail using
the method of tissue cultured irradiated organs and discs.
Wild strains (Oregon, Canton-S, Koehi, and Samarkand) and several
eye-color mutants (bw, w, v, en, v bw, en-ew Bar, bar-3 and DpfIn(3L)P,In(3R)C, Sbel(3)e of D. melanogaster were used as material. Third-instar
larvae (about 80 hours old after hatching at 25°C) grown under sterile
conditions were irradiated with various doses of X-rays (160 Kvp, 25 rnA,
Imm Al filter, 370 rfmin, at the distance of 30 em). They were dissected,
and the various organs and tissues were removed under a binocular
microscope in a sterilized glass chamber. They were cultured in asynthetic medium, and the sensitivity for irradiation was determined byobserving their growth and differentiation. Some of the results obtained
from a wild strain (Oregon) are shown in Table 1.
When the irradiated (0-20 Kr) eye discs were cultured together with
the normal cephalic complexes (ten bodies), they showed pronounced
growth and differentiation as in normal eye discs. But after high doses

RESEARCHES CARRIED OUT IN 1957

76

Table 1. Growth and differentiation rate of the irradiated eye
discs in tissue culture.
Organs and discs culturedI,

G.* and D.*
rate----------

o 0.5

1

Doses (Kr)3 5 10 15 20 25-- - - . - . - -N.* eye disc+I.*C.c.*

G

+
+

+
+

+
+

+

+

+

+

±

D

I. eye disc+N.C.C.

G
D

+
+

+
+

+
+

+
+

+
+

+
+

+
+

±
±

I. eye disc + I.C.C.

G
D

+
+

+
+

+
+

+

+

+

+

._--------------_.-._-

Culture medium

G
D

+
+

+

+

+

+
+

+
+

Extracted metamorphic
hormone

G

+
+

+
+

+
+

+
+

+
+

D

* G.=Growth, D.=Differentiation, c.C.=Cephalic complex (ten bodies), N.=
Normal, I. = Irradiate.

(25 Kr), neither growth nor differentiation were observed.
Otherwise, when normal eye discs were cultured together with the irradiated (3 Kr) cephalic complexes (ten bodies), the eye discs showedpronounced growth, but no differentiation. Since the cephalic complex is
known to exert hormonal control on growth and metamorphosis, it seems
that the normal differentiation mechanism of organs and discs of the
cephalic complex was virtually destroyed after low doses (3 Kr). Hence,
one might suspect that bar-3 mutant which showed high radiosensitivity
in whole body irradiation test of thirteen strains used would be due tohigher sensitivity to radiation damage of the cephalic complex than otherstrains.
Furthermore, the metamorphic hormones already secreted from thecephalic complexes were not influenced even after high doses (25 Kr).
The result obtained in this experiment shows that the delay in pupationand the decrease in imagination rate are due to radiation damage of the
cephalic complex.

51.
1.

Structure and Function of Pleiotropic Genes in the Silkworm
An Experiment to Break up Em, EKp and ENe genes into
Separate Components by X-rays (a Preliminary Note)(By Yataro T AZIMA and Eiichi INAGAKI)

In genetically well analysed organisms some genes have been known toRESEARCHES CARRIED OUT IN 1957

77

control so diverse characters that no relation between them would seemto exist. For instance EKP controls the appearance of extra legs andextra markings in different segments of the silkworm. The question
arises if it is possible to divide the gene into some unit components which
control the seemingly different characters. Is there any change in the
manifestation of each component when they are separated or when they
are brought together under a new sequence? In order to get an answer
to these problems, the present study was initiated.
Efill (extra crescent markings on the fourth segment and extra legs on
the fourth and fifth segments), EKP (extra crescents on the sixth segment
and extra legs on the fifth segment) and ENc (no crescents on the fifth
segment and extra legs on the same segment) were used as materials.
X-ray irradiation was administered to the males of those mutants at thepupal stage. Irradiated individuals were crossed to untreated partners ofnormal strains (with crescents on the fifth segment) after emergence. F 1
offspring was examined carefully for dislocation of crescent markings andextra legs. Several exceptional individuals were discovered with respect
to both characters and classified into E, ECa, ED, EO, ... etc. according
to their phenotypes, the corresponding mutant types being already known(Table 1). Genetic analysis has not yet been carried out.
Table 1.

Aberration induced by X-rays in Efll, EKP and ENC

Cross

Aberrations induced+/+¥xEfll/+(X-rayed) ~+/+¥ xEKP/EKP (X-rayed) ~E, EGa, ED, EKP, EN, ENC and new types
ECa, ENc, homo EKp and a new aberrantwith regard to abdominal leg
EKp+/+¥xENC/+(X-rayed) ~Most of the mutations occurred in the direction from complicated types
to simpler ones, but in very few cases the reverse was true. The
frequency of appearance of the aberrants was far higher than would havebeen expected from a single gene locus which indicates that the E-allelicgene is not a single unit locus but an aggregate of small unit loci which
control different characters.
52.

Silkworm Larvae with Extra-abdominal Legs Caused
by a Chromosome Fragment(By Mitsuo TSUJITA)

Female pupae of the genotype ElTEKP/E"EKP were irradiated with X-

78

RESEARCHES CARRIED OUT IN 1957

rays of 8000 r, and the imagines were mated to normal males. The larvae
with supernumeral abdominal legs caused by a chromosome fragment
were found in the offspring of this cross. Small extra-abdominal legs
were produced on the 4th and 5th abdominal segments of the mutant
larvae. Those on the 4th segment were often missing.
As the phenotypic manifestation resembles that of E, the gene symbol
E' was given to this mutant.
From the cross + x E' or E' x E' the E' larvae segregate as shown in
the following tables:
Table 1.
Hatching rate

+

92.6

1158

+xE'
Total

363

-_ _ - ...

Table 2.

I

1521------------------

E' x E'
Total

Hatching rate

+

87.8

823

1715

892

The number of the E' larvae in Tables 1 and 2 is less than that whichis theoretically expected to segregate. The E' homozygous individuals
could not be ascertained because they are hardly different from theheterozygotes. To examine the linkage relation between E' in question
and EGa belonging to the E region of chromosome VI, cross experiments
were carried out.
From the cross E'/EGa X + and from its reci procal four types +, EGa,
E' and E' EGa segregated as shown in the following table.
Table

E'EGax +

+

EGa

E'

E'EGa

Total

94

63

64

16

237

-------.---

Total of two batches

_ ..

--

.. _ - - - - - - - -- - - - - - - ----Table

+ xE'EGa

+

EGa

850

377

E'

---

,E'EGa I Total------c--- -------

Total of two batches

I

I

568

134

1929

- - - - - - - - - " - _.. _ - - -From the experimental results shown above, it is evident that E' does
not link to chromosome VI.

RESEARCHES CARRIED OUT IN 1957

79

According to my cytological observation, a very small chromosome which
is probably derived from breakage of chromosome VI could be observedat the maturation division of the first spermatocyte of the E' individuals.
It seems that the characteristic trait of E', supernumerary abdominal legs,
is produced by the genetic constitution of E on the chromosome fragment.
Furthermore, it may be inferred that by the treatment with X-rays
this fragment was formed as a result of breakage of chromosome VI, at
the end of which ER EKp loci are located. As stated above, mutant
larvae have two supernumerary abdominal legs but they have not theextra crescent patterns on the dorsal side of the 4th segment. Why thesepatterns do not appear?
It is possible that in spite of the presence of £II locus the extra crescentpatterns are not produced owing to a position effect. On the contrary, it
may be due to a defective structure of the EH locus. On this point
further studies are required.
The attachment of spindle fibres on the chromosome fragment should
be taken into consideration.
f

53.

Effect of Chronic Gamma Irradiation on the Breeding
Behavior of Mice at Different Developmental Stages(By Tsutomu SUGAHARA, Kiyosi TUTIKAWA, Yoshihiko SUGIURA,
and Tomizo TANAKA)

As previously reported (Annual Report No.7), the effect of chronic irradiation on the breeding behavior was different according to the developmental stages and sex of the irradiated mice. A similar experiment wasundertaken on a larger scale with respect to the number of animals andthe number of developmental stages in order to obtain more detailed information about the difference between the effects.
Mice of the CBA strain kept in aluminum cages were subjected tochronic gamma irradiation. Upon completion of irradiations, all animals
were mated with mice of NH strain by placing one male with one or two
females. Irradiations were made from the r-ray source of 60CO in a
specially designed irradiation room, for 22 hours per day and 5 days per
week. A total dose of 450 r was reached in about 80 days in the firstseries, and that of 366 r in about 70 days in the second series.
The treated mice were divided into four groups, A, B, C, and D according to their developmental stages during irradiation. Group A consisted of animals receiving radiation from fertilization, through birth,
until the age of 50 to 60 days. Group B was composed entirely of adultanimals. In group C and D, the irradiation started with birth and weaning

RESEARCHES CARRIED OUT IN 195780

respectively.
The results of matings examined during three months after irradiation
are summarized in Table 1.
Table l.
==-,--_"":",,:,,,==-:

group

Dose

Breeding behavior during three months after irradiation.
.. _---_Mean
No. ofSex% ofirradiatedSterility
litterSex
ratio
controlsize
animal-------------_._--.. _ - - -------------------

A

if-

B

if-

C

if-

D

if-

A

if-

B

if-

450r

366r

88-

8-

88-

8-

.-------_._, ..

0
100
40
8.3
15.4
88.9
88.9
61.5

5.55

81.7

61.0

4.42
4.83
5.2

65.2
71.2
77.1

71.7
56.3
62.5

3.7

54.6

43.3

10
7
9
6

10
100
0
16

5.32

73.8

48.0

4.25
4.91

62.6
72.3

65.6
50.0

6.78

100

54.5

,._-

Control- - - - - - - - - - - - - - - - _._------_

31
34
20
12
13
18
9
13

23
..

- - - - _ ----.
...•

_-~

Sterility and litter size showed similar tendencies among different groups
in each sex. In the female, sterility percentage and litter size reduction
became less pronounced in the order of the age of irradiated animals.
In the male, opposite results were obtained on these points with the exception on group D. As for sex ratio, which has recently been regarded
as one of the most useful means to estimate the genetic effects of radiation on man, the results were rather controversial, though in the caseof irradiated males of group A a statistically significant increase wasobserved in the proportion of males. With regard to the changes ofbreeding behavior following irradiation, the males seemed to be able torecover from the damage, and the recovery was earlier for younger
males. On the other hand, the females, seemed to become permanently
sterile sooner or later, especially the younger females which were sterile
from the beginning.
Recessive mutations at three loci of the double recessive NH mice were
not observed in F 1 consisting of 819 animals.

RESEARCHES CARRIED OUT IN 1957

81

54. Biochemical Studies on the Radioprotective
Effect of Cysteamine(By Goro YAMAMOTO, Saburo NAWA and Tsutomu SUGAHARA)

It has recently become known that in mammals cysteamine is one of
the most effective protective agents against lethal effect of radiation. In
order to clarify the mechanism of its protective effect, disturbance of
nucleic acid metabolism induced by X-irradiation was studied in the spleen
and thymus of the mouse, some partly with and partly without cysteamine
administration before irradiation.
Female mice of the NCF 1 strain (NH x CBA) were used for the experiment. Groups of three or four animals were sacrificed at one time an@dissected to obtain the spleen and thymus. Each organ was weighed immediately and nucleic acids were extracted by the Schneider method.
DNA content determined photometrically by diphenylamine color reaction,
and RNA content by orthin color reaction.
Total DNA content in the spleen and thymus as well as their weight
were markedly decreased when the whole body was irradiated, even withrelatively low doses of X-ray as Table 1 shows.
Table 1.

Total DNA content and weight of spleen and thymus,
24 hours after X-irradiation.
Spleen

i

Or
68r
136r
300r

ThymusDNA content in %Weight in %DNA content in %Weight in %

100
55.4
38.2
22.9

100
56.2
43.5
36.5

100
61.8
35.2
17.1

100
59.4
43.4
28.2

-----------------

Concerning the effect of cysteamine, DNA and RNA content in theorgans were studied one, two and four days after irradiation with 300 r.
Animals examined were divided into four groups; 1) those injected intraperitioneally with a dose of 0.15 mg/body weight of cysteamine thirty
minutes before X-irradiation, 2) those injected at the same time intraperitoneally with 0.5 cc of physiological saline, 3) those injected intraperitoneally with the same dose of cysteamine as the first group but without
X-irradiation, 4) untreated controls. The mice of group 1) could survive
even when exposed to a lethal dose of 800 r. The weights, total DNA
and RNA contents of spleen and thymus of the treated animals as ex-

82

RESEARCHES CARRIED OUT IN 1957

pressed in per cent in relation to controls are shown in Figs. 1 and 2.
%
100

Cystemine +X-ray (300r)
Salin +X-ray (300r)
Cysteamune
Organ weight
A

DNA
RNA

50
40

30
20

/0

/0

o

J

4 daysFig. 1. Changes in weight, total
DNA and RNA contents of thethymus after treatments.

o

2

3

Fig. 2. Changes in weight, total
DNA and RNA contents of thespleen after treatments.

In both organs reduction of the three value at one and two days after
irradiation was quite similar in both cysteamine-treated and cysteamineuntreated group. However, at four days after irradiation the results weredifferent depending on the presence or absence of cysteamine treatment.
Recovery, especially in RNA content, was seen in the treated, but not in
the untreated group. In addition, it should be emphasized that both DNA
and RNA content were reduced by the administration of cysteamine
alone. Cysteamine itself seems to be a toxic substance to nucleic acidmetabolism and yet able to protect the metabolism from the radiation
damage by enhancing its recovery. Such a contradictory phenomenon
was also observed in body weight. The reduction of body weight justafter X-irradiation was more marked in the cysteamine-treated groupthan in the untreated one. After four days, on the contrary, the recovery
of body weight was striking in the cysteamine-treated group even whenirradiated with a dose lethal to the untreated.

55.

Melanotic Tumors Induced in Drosophila melanogasterby Ionizing Irradiation(By Toshifumi TAIRA and Toshiteru MORITA)

Melanotic tumors are induced easily by ionizing irradiation in larval

RESEARCHES CARRIED OUT IN 1957

83

tissues of D. melanogaster if early larvae are exposed to radiation with
800 and more roentogens. The frequency of induced melanotic tumor
differs according to mutants or dosages. In the former case, it seems todepend on differences in gene constitution in respect to metabolism. In
the latter case, it may be due to the ionizing intensity of the agent. The
results are shown in Table l.
The isogenic line of Hn r3 , one of the eye color mutants, shows thehighest frequency of induced melanotic tumors. Therefore, it was used
to analyze developmentally and biochemically the formation-mechanism of
melanotic tumors induced by ionizing irradiation. The effects of trypotophan and cysteamine added to a normal medium on the frequency of induced melanotic tumors were examined. The results are given in Table 2.
By adding tryptophan, the frequency of induced melanotic tumors isdistinctly increased, but it is conversely decreased by adding cysteamine
to the culture medium.
Although constituents between irradiated and non-irradiated groups of
prepupae were made a comparative study chromatographically, there are
not any distinct differences among the free amino acids besides excessive
phenylalanine in the irradiated group without melanotic tumors.
Table 1.

Frequencies of the induced melanotic tumors by ionizing
irradiation in D. melanogaster strains.

Dosage(Kr)

No. of treated 1st larvae

No. ofpupation

No. of pupae
with melanotic tumors% of induced

StrainOregon-2

0
3
5
8

300
680
550
603

288
453
383
360

0
10
16
11

0
2.2
4.2
3.1

0
3
5
8

300
150
578
315

283
84
224
71

0
10
18
7

0
11.9
8.1
9.9

0
3
5
8

102
78
50
123

82
55
41
6

0
16
7
1

0
29.1
17.1
16.6*

ell

melanotic
tumors-----------

Hn T3

*)

Decrease of frequency with higher dosage seems to be due to increase oflarval lethality.

84

RESEARCHES CARRIED OUT IN 1957
Table 2. Frequencies of melanotic tumors induced by ionizing irradiation
after adding tryptophan and cysteamine to the culture medium (Hn ro , 3 Kr).

Non-irradiatedControl
Tryptophan(5% )**
Cysteamine(20 JLgjml)

IrradiatedControl
Tryptophan(5%)**
Cysteamine(20JLgjml)

No. oftreated 1stlarvaeNo. ofpupation

No. of pupae
with melanotic tumors

300

253

0

0

300

233

0

0

300

155

0

0

500

488

78

16.0

550

500

144

28.8

330

141

9

6.4

% of induced
melanotic
tumors--------**J

Tryptophan is added at 5 per cent of a given yeast powder to the medium.

56.

Differential Sensitivity to Xray Irradiation of Maleand Female Day-old Chicks(By Takatada KAWAHARA)A study of mortality in response to whole body exposure to X-rays has
been carried out in day-old chicks of two breeds, namely White Leghorns(WL) and Barred Plymouth Rocks (BPR) and their reciprocal F 1 hybrids.
The irradiation was made under the following conditions: single dose of
800 r, at 180 KV, 25 rnA tube current, added filter 0.3 mm copper and
0.5 mm aluminum, in air at the target distance of 50 cm, intensity 82.5 r!
min. All experimental birds were reared in electric brooders and were
fed a commercial chick starting mash. Observations were made up to 20
days after irradiation. Lethality directly due to irradiation by X-rays apparently occurred within 20 days after treatment, afterwards the increasein mortality was negligible compared with that of non-treated controls.
The percent mortality and its statistical analysis are given in Table l.
In general, the males were more radiosensitive than the females. Differences between sexes of pure-breds were significant (X 2 = 11.64, P.01),
however, those in hybrids approached the significance level (x 2 =3.75, .10
.05).

<

>P>

85

RESEARCHES CARRIED OUT IN 1957
Table l.

Mortality of various mating types exposed to irradiation
by a single dose of 800r.
Breed or
cross

Sex

No.
testedDead birds~ ~ - - - -

No.

%

x2

- -----------WL
BPR
Treated (SOOr)
WLif.x BPR'CS'
BPRif. x WL 2)

'CS'
if.
'CS'
if.
'CS'
if.
'CS'
if.

53
47
15
16
47
65
55
43

'CS'
if.
'CS'
if.
'CS'

20
16
17
12
10
9

46
29
15
12
40
44
41
29
~ - - - - _ . _ . -WL
BPR
Non-treated
WLif.x BPR'CS'
BPRif.x WL'CS'

if.
'CS'
if.

11

12

86.8}
61. 7 11.22**}100.0
11.64**
75.0} 4.53*
85.1 } 4.41*
67.7
74.6} 0.59
67.4

3.75

~---------

1

0
0
0
0
10.0

0
0
0

0
0
0

0
0
0
0

I

-----------*J**J

Significant at the 5% level.
Significant at the 1% level.

57.

A New Spacing in the Wide Angle Xray Diffraction
Pattern of Dried Striated Muscle(By Tsutomu SUGAHARA, Masayoshi OZAWA*,and Nayao UESUMI*)

Striated muscle dried at its normal length has been reported to show
typical wide angle X-ray fiber diagram with a main meridian and a equatorial spacing around 5.1A and 9.6A analogous to that of a-keratin. Dried
actomyosin film, a contractile protein of the muscle, was reported to showa wide angle diagram similar to that of the dried muscle. Based on thosediagrams ASTBURY presented his famous molecular model of a-keratin
type. Very rarely, however, a number of spots, as are usually found in
LOTMAR-PICKEN diagrams, have been obtained, which PAULING and COREY*

Mie Prefectural University School of Medicine.

86

RESEARCHES CARRIED OUT IN 1957

believed to support their concept of a helical configuration of fibrous protein. While examining the X-ray diffraction pattern of dried striatedmuscle by an automatic recording X-ray diffraction apparatus, we foundthe diffraction peak to be around 5.9A which had never been noticed in
the above mentioned diagrams. The same spacing was observed in dried
and powdered muscles as well as in dried actomyosin gels.
Muscle specimens were taken from psoas muscle of the rabbit. Actomyosin was prepared from rabbit muscle by the method of SZENT-GYORGY
with repeated purifications. Actomyosin gel formed in distilled water waswashed several times to remove salts completely, for even a small quantity of residual salts could disturb the X-ray pattern with their diffraction
peaks. All diffractions were done by a NORELCO automatic recording
X-ray diffraction apparatus operating at 40 kVp with Ni-filtered copper Karadiation.
Some tracings of thepatterns obtained from598.0.
28=5° to 20° are reproduced in Figure l.
Curve a) was obtained
from a resting muscle
dried at normal length;
the spacings perpendicular to the fiber axis
are shown. Curve b) was
obtained from a muscle
in rigor mortis dried at
its original length in thebody; curve c) represents a resting muscle
dried and powdered;curve d) correspond todried
and powdered
muscle in rigor mortis;
curve e) was obtained
from actomyosin gel.
4·5
9101112
Sharp peaks around 5.9A
17 A
spacing were observedin all curves. They were
Fig. 1. Tracings of the X-ray diffraction patternof different height acof dried muscle from 29=5° to 20°. Explanation
cording to the various
seen in the text
specimens. In the case

RESEARCHES CARRIED OUT IN 1975

87of the dried muscle fixed at its normal length the peak was quite prominent and sharp, and more so in rigor mortis. Slight variation in
spacings and fairly large differences in their height and half value widthappeared to occur according to the physiological condition of the muscle;
their significance is not yet clear. A comparison of the curves obtainedin two directions, parallel and perpendicular to the fiber axis, showed thatthe spacing was oriented mainly perpendicularly to the axis.
Further, a peak around 5.1A, usually recorded by conventional film
method, has not been found. In some cases, there appeared to be one or
two other sharp peaks especially in the region of 3.7A. However, as theirincidence was not consistent, the. experiments are now being repeated.
Since the above data were markedly different from those of previous
workers, the origin of the curves should be taken into consideration.
Anorganic substances, if cristallized in the muscle by loss of water, cangive sharp diffraction peaks characteristic of their crystal lattice, which
are classified and tabulated in Hanawalt's Table.
Substances showing about 5.9A diffraction peak can hardly exist in themuscle according to the Table. It may be possible that the diagram wasoriginated mainly from the non-contractile component of the muscle, forexample the muscle membrane. The fact that the actomyosin gel showeda peak around 5.9A, though its height was very low, seems us to beagainst this possibility.
In conclusion, while these data are not sufficient to warrant a new pro'posal of the molecular configration of muscle protein, they point out that
5.1A spacing is not suitable for its elucidation.

J.

RADIATION GENETICS IN PLANTS

58. Effect of X- and r-irradiation on Chromosome Aberrations
in Vicia faba
(By M. NEZU and S. MATSUMURA)

The beans were soaked in tap water for 48 hours (16 c C) and immediately after the removal of the seed coats irradiated by X- and r-rays inwater. The dose of both rays was measured by ferrous sulphate dosimeter (FRICKE'S method) in order to secure irradiation at the same dose(600 r) for the same time. After irradiation the beans were kept in adark room at 20°C and their root-tips were fixed 24 and 30 hours later,
and then treated by FEULGEN squash method. The observations of chromosomal aberrations were arbitrarily limited to cells in early telophase

88

RESEARCHES CARRIED OUT IN 1957
Table 1. Comparison of chromosome aberrations of X- andr-rays induced in Vieia faba roots.
Total no.
of cells
observed

SectionControl 24 hr

X-24 "

r- 24 "
X-30"

r- 30 "

400
300
300
650
650

Percentage ofI abnormal
I cells (%)

Average no. per cell

I

0.0
0.669
0.306
0.300
0.184

I

I

I
I

2.3
70.3
57.0
62.6
39.4

Bridges!

Fragments

Averageof breaks
per cell

0.023
0.716
0.582
0.775
0.425

0.023
1.170
0.760
0.938
0.538

just before the formation of nuclear membranes. In this stage, the chromosomes are grouped at both poles, and are stained as strongly as at
anaphase or metaphase. We could easily observe bridges and fragments
and ascertain their relationships. The percentage of cells with abnormal
chromosomes was about 70% and 57% 24 hours after X- and r-irradiation
respectively, while after 30 hours, it was reduced to 63% and 39% respectively. This indicates that X-irradiation is about 1.2-1.6 times more
effective than r-irradiation. At the former, the average number of bridges
per cell is 2.2 times more after 24 hours than at the latter, and 1.6 times
more after 30 hours, while in respect to fragments it is 1.2 and 1.8 times
more respectively.
It is assumed that the breakage occurred when the chromosomes were
not yet split, so we estimated the number of breaks as follows: a fragment or a bridge depends on one chromosome break, and when there
are more bridges than fragments the number of breakages is estimatedfrom the number of bridges and vice versa. The average number ofbreaks per cell, after 24 hours, was 1.17 in X- and 0.76 in r-irradiation,
while after 30 hours it was 0.938 and 0.538, respectively. As to chromosome breakages, X-irradiation produced them about 1.7 times as many as
r-irradiation.

59. Effects of Temperature and Irradiation Time upon
Mutations Induced by Radiations(By Seiji MATSUMURA)

Dormant seeds of Triticum monococcum flavescens were exposed to Xand r-rays at the dosage 10 and 20 kr. The growth of seedlings, thesingle-spike fertility and chromosome aberrations of treated plants (Xl)RESEARCHES CARRIED OUT IN 195789

and the chlorophyll mutations in X2 were compared for acute and chronic
irradiation. At acute irradiation with X- and r-rays treatment was given
either immediately before sowing or the irradiated seeds were kept for
30 days in storage at room temperature (about 20°C) or at 5°C. At chronic r-irradiation with 60CO the treatment lasted 54 days. Also, the effectof S-radiation by 32p was examined for comparison.
The data are shown in Table 1. The relation between the inhibitionof seedling growth and dosage, temperature in storage and irradiation
time coincides roughly with the relation between the percentage of induced sterility and all those conditions. X- and r-irradiations were far more
effective at 20 kr than at 10 kr. In the case of 30 day storage, r-rays inhibited the growth of seedlings and reduced the fertility more than Xrays, while irradiation applied just before sowing showed the reverse
relation. It was found further, especially with r-rays, that low temperature had the strongest inhibitory effect. At 10 kr acute r-irradiation was
Table 1. Genetic effects of ionizing radiation in Triticum monococcum.

30 day storage
at room temperature

30 day storageat 5°C

Dosage(kr)

Length ofseedlings*(cm)

Fertility of
spikes inXl (%)

Chromosome
aberrations
in Xl (%)

Gene
mutations inControl

17.69(11.08)

74.62

0.00

0.0

X-10
X-20

14.99
13.53
8.96
6.79
13.81
11.63
9.73
3.45
10.71
4.29
12.00
6.75
14.69
5.37
(12.66)(14.14)

81.82
11.59
60.32
32.78
61.95
33.53
60.58
8.34
62.32
34.65
64.38
40.10
66.45
15.47
68.60
79.28

0.00
19.05
14.29
25.00
0.00
54.17
4.08
40.00
7.50
20.00
10.00
38.46
4.08
28.57
0.00
2.50

0.0
16.7
5.7
11.1
2.3
14.3
9.1
33.3
6.1
13.3
5.3
5.6
5.3
50.0
1.9
4.1

1r-r1

lO

20

X-10
X-20
r- 10

r- 20
X-lO

Ac~te
~rradia- 1
hon Just

X~20

before sowing
Chronic
irradiation
32p

r- lO
r- 20
r-10

{

{

r- 20
tHO

fl-20

X2 (%)---'--------'------~.--------* X- and r-irradiated seeds were sown October 25th and the seedlings were
measured 27 days after sowing. ( ) Sown October 27th and measured 25days after sowing.

RESEARCHES CARRIED OUT IN 1957

90

more effective than the chronic one, while at 20 kr the reverse relation
was observed.
The frequency of ears with chromosome aberrations in XI·plants was
strikingly higher at 20 kr than at 10 kr. In most of the cases of induced
chromosome aberrations ®+5 n , often 6n +2 r , ®+4 n , ®+®+3 n or ®
+4 n +2 r and seldom 1m +5 n +1 r or asynaptic 14r have been observed.
The effect of r-ray was generally stronger than that of X-rays. Also, irradiation just before sowing and 30 day storage at low temperature produced more chromosome aberrations than storage at room temperature
after irradiation. On the other hand, the effect of chronic r-irradiation
was less. This must be due to the longer time of chronic irradiation,
because two-hit aberrations, such as translocation, are limited in time.
The frequency of head progenies with chlorophyll mutations in the X2 generation increased with the increase of radiation dosage. Because of
the small number of observed head progenies, due to a lower survivalrate, the results with 20 kr irradiation were unsufficient. But they wereroughly in accord with the observations of chromosome aberrations.
The effects of l3-irradiation were unexpectedly slight. It was foundfrom another experiment with seed absorption of 32P-solution that theactual dosage of l3-rays was very low.

60.

Crosses between Various X-ray Induced Recessive
Mutants in Wheat(By Taro FUJII)

Several mutant strains of Triticum monococcum fiavescens, namely
chlorina, basi-viridis II, virido-albina, slender and irregular-ear were crossed with each other. All of these mutations were in crosses with normals
simple recessives. In F2 dihybrid segregation was observed and double
recessive segregants were obtained.
Chlorophyll content of double recessive plants from the cross viridoalbina x chlorina was slightly decreased in the seedling stage, comparedwith virido-albina itself. When they were placed in the phytotron (20°C,
80% relative humidity), their leaves gradually turned to light green andincreased the chlorophyll content, until its amount was restored to thechlorina level, but a further increase was never observed. Seedlings
obtained from the double recessive plants from the cross virido-albina x
basi-viridis II had no chlorophyll just like albina and died out a half monthafter germination, even in the phytotron.
Double recessive plants in a cross virido-albina x slender or irregular-

RESEARCHES CARRIED OUT IN 1957

91

ear were in the seedling stage similar as to chlorophyll to virido-albina
itself. When both were placed in the phytotron, their chlorophyll content
gradually became restored and about a month later was as that of thenormals.
From these experiments it follows that the chlorophyll content of viridoalbina could be recovered in the cross combinations with slender and irregular-ear which have a normal chlorophyll content. But the viridoalbina gene was hypostatic to the chlorina gene and the same behavior
was shown by the double recessive plants between basi-viridis II and chlorina (Ann. Rep. No.7). Albina-like plants obtained from the cross between virido-albina and basi-viridis II must have been genetically different
from albina mutants but they behaved like the latter.

61.

Radiosensitivity in Plants(By Taro FUJII)

Determination of radiosensitivity in plants, such as LD 50, is important
in radiobiological studies and in plant breeding by radiation. Radiosensitivity is quite different in different plants. A comparison of radiosensitivity in many plants, especially in related plants of various polyploidseries, were carried out in 1957.
A. Experiments in crop plants
Dry dormant seeds of several crop plants were irradiated by r-rays
from 60CO source. Dose ranges were 10-100 kr, and radiosensitivity was
examined by the decrease of germination rate. Some of the results areshown in Table 1.
Germination rate in most of these plants gradually decreased with theincrease of doses, but that of Cryptotaenia, Lilium or Oryza abruptly decreased to 0 at a certain dosage. Germination ability up to the appearance of cotyledons developed in several species, for instance, Glycine
or Raphanus, not much even at 100 kr, but most of the seedlings died
immediately after the germination, and only a few plants could survive.
Radio sensitivity must therefore be determined by the percentage of seedlings which had developed 2 or 3 leaves.
From this experiment, the genera Cosmos and Brassica were most resistant to radiation, while Cryptotaenia and Lilium were most sensitive.
B. Experiments in Triticum and Aegilops
Differences of sensitivity between di-, tetra- and hexaploid plants werecompared in several species and varieties of Triticum and Aegilops. Dry

RESEARCHES CARRIED OUT IN 1957

92

Table 1. Germination rate in several crop plants.
MaterialCont.

Cosmos bipinnatus
Capsicum annuum
Vinca major
Cryptotaenia canadensis
Impatiens balsamina
Glycine max
Arachis hypogaea
Vicia faba
Raphanus sativus
Brassica campestris
Fagopyrum esculentum
Lilium philippinense
Zea mays
Oryza sativa

70%
50
42
82
81
90
88
99
60
97
97
42
89
81

19
20
0
66
84
81
73
47
100
91
0
91
81

40 kr

70kr

100 kr

64%
0
0
0
43
78
88
15
48
100
75
0
89
67

56%
0
0
0
55
1
42
4
34
98
16
0
61
0

43%
0
0
0
0
0
2
1
26
98
0
0
26
0

dormant seeds were used for this examination which were irradiated from
20 to 70 kr by r-ray. Radiosensitivity was determined by the decreaseof germination rate and the average length of the seedlings. Genus
Aegilops showed more tolerance to radiation than genus Triticum, in
tetraploid species of Aegilops the germination rate remained even at 40 kr,
compared with that of the control, while that of the tetraploid Triticum
dicoccum showed a marked decrease at 30 kr. A part of the results isshown in Table 2. Diploid species of Triticum and Aegilops were theTable 2. Germination rate in cereals.
Material
T. aegilopoides
T, monococcum
T, dicoccum
T. durum
T, Spelta
T, vulgare
Ae. cylindrica
Ae. uniaristataAe, squarrosa
Ae. ventricosa

2nGenome

14 AA

"

"

"

"

"

"

28
14
14
28

CCDD
MuMu

28 AABB
42 AABBDD

DD
DDMvMv

Cont.

20kr

30 kr

40kr

30.0
61.3
62,5
50.0
71.3
75.0

6.3
47,5
73.4
66.3
72.2
62.5

0,0
38,0
33,8
55,0
76,3
50.0

0.0
12.5
21.3
42.5
50,0
42.5

0,0
21.3
0.0
15.0
10.0
15.0

86.3
58.2
72.5
61.3

82.5
71.3
66.3
77.2

78,8
81.3
65.0
63.8

56.3
12.5
32.5
70.0

0,0
0,0
0.0
0.0

i

70 kr

93

RESEARCHES CARRIED OUT IN 1957

most sensitive to radiation, and the hexaploid Triticum species were notmore resistant than the tetraploid ones.
Differences of sensitivity were also observed in different varieties (orspecies) of a species (or groups). These facts show that radiosensitivity
depends not only on the kind and number of genomes but also on thekind of alleles present.

62. Radiosensitivity of Several Commercial Varieties andUseful Mutants Induced by X-rays in Tobacco(By S. MATSUMURA and T. FUJII)

Dry seeds of 5 commercial varieties, Bright Yellow, Dixie Bright 101,
Golden Wilt, White Burley and Matsukawa, were exposed to the same
dosage (30 kr) of X- and r-rays. There was a difference in radiosensitivity among those varieties. The germination rate of treated seeds markedly decreased in Bright Yellow, Dixie Bright 101 and Matsukawa, especiallyat r-irradiation. In these commercial varieties the germination and thegrowth of seedlings were more delayed.
The mutant "early" of the variety Bright Yellow was irradiated in1956 with X-rays at 30 kr. In the X2 -generation many recessive mutants
were observed, such as abnormal flowers, narrow leaves, etc.
" Early" (No.6), "pubescent" (No. 12) and "yellowish green" (No.
Table 1.

Yield in kg and value in Yen of leaves of X-ray mutantsof flue-cured tobacco (1957).
-_._-

Strains% of drymatterYield in kg
per Tan*Bright Yellow
6-2-2A
6-2-2B
6-3
12-2A
12-2B
12-2C
13-2A
13-2B
13-2C

16.5
16.2
15.7
19.0
15.8
15.6
14.5
15.1
15.3
15.5

136
115
123
143
135
155
149
133
129
120

I

~~-----* Tan=ca. 0.1 ha.

Value in Yen
per kgAverage
grade

Return per
Tan* in Yen

222
222
229
226
312
287
311
281
269
323

5.25
5.25
5.15
5.20
3.76
4.21
3.76
4.31
4.51
3.54

30,20625,52628,52632,172
42,08444,453
46,33037,44034,70638,755RESEARCHES CARRIED OUT IN 1957

9413), mutants of Bright Yellow, were of good quality and valuable forthe improvement of tobacco. Table 1 shows the results of comparative
experiments with Bright Yellow as standard in the field of our institutein 1957. From this table No. 12-2 is the best and No. 13-2 is next to it.
Both strains show clearly better results than Bright Yellow. Simultaneously similar comparative experiments with these strains were carried out
on a large scale in several Tobacco Experiment Stations of Japan Monopoly Corporation (Mito, Utsunomiya, Hatano, Misima, Iwata and Hyogo).
There was a difference among the experimental fields. Generally No. 12-2 and No. 13-2 again showed better results, especially in the southern
districts. The latter strain was slightly better than the former. No.
12-2 is not earlier than Bright Yellow and its flue-cured leaves, especiallylower ones, are thick and rough, and have hard texture and low waterabsorptiveness. No. 13-2 is a few days later than Bright Yellow andhas slightly more leaves, of which the upper ones are thin and shiny andhave a fine texture and high absorptiveness. Both strains are of goodquality and very promising in the southern districts.

63.

Induction of Bud-sports by X- and r-rays(By S. MATSUMURA and T. FUJII)In Cinnamomum camphora seedlings one year old and cuttings were
exposed to X-rays (180 KVP, 3 rnA, 240 rfmin) and r-rays (60CO, 0.6-1
kr/hr) at the dosage 1, 2, 4 and 6 kr. The number of successful planting
and sprouting per seedling or cutting shoot decreased in proportion to theincrease of radiation dosage. X-rays were more effective in this respect
than r-rays (Tab. 1). Also, the frequency of plants with abnormal ordeformed leaves increased roughly with the increase of dosage.
In the variety Norin No. 1 of sweet potatoes (Ipomea batatas) eachroot-tuber was longitudinally cut in 2 pieces. Each half was subjected to
X-ray (180 KVP, 25 rnA, 154 rfmin) and r-ray treatments (20 days by 60CO)
at 2.5, 5, 10 and 15 kr. The other half was untreated and used as control. X-rays effectively inhibited sprouting, while the effect of r-rays
was not so much. When X-irradiation was applied the frequency of
sprouted tuber-halves and the number of sprouts per tuber-half decreased
with the increase of dosage. When treated with X-and r-rays, sprouting
was always markedly delayed, especially at higher dosage. After planting of these sprouts, white, white-stripe, deep color and oblong shapewere often found as chimera in new root-tubers.

95

RESEARCHES CARRIED OUT IN 1957
Table 1.

Effect of radiations on sprouting in Cinnamomum camphora.

Tre~~~ents-(DO;)"-=--I-C~~~. - - 1 k;=-2kr-~k~-6 kr
No. of seedlingsprouted (%)
No. of sprout
per seedling
No. of abnormal
seedling (%)

X-ray

97

83

70

9

7

3.2

3.0

3.1

2.4

2.0

3
(3.1)

8
(9.6)

12
(17.1)

3(33.3)

1(14.3)

96

98

94

91

50

2.8

2.9

3.4

2.9

2.6

1
(1.0)

7
(7.1)

7
(7.4)

10
(10.9)

13(26.0)

86.96 77.55

77.51

38.00

17.65

1. 4

1.5

1.2

1.2

1.3

62.50

59.18

33.33

1.5

1.2

1.1

Seedling
No. of seedlingsprouted (%)
No. of sprout
per seedling
No. of abnormal
seedling (%)

r-ray_ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _c

No. of seedling!
sprouted (%)
No. of sprout
per seedling

I

X-ray

_

Cutting
r-ray
----

---

No. of seedlingsprouted (%)
No. of sprout
per seedling------- - - - - - - - - -- - - - - - - - - -64. Induced Polygenic Mutations in Rice
(By Hiko-Ichi aKA and liro HAYASHI*)

From seeds of a pure line of rice, a control and two X-rayed (6,000 r
and 12,000 r) plots were prepared. They were grown in bulk for four generations. Plants with any recognizable change (sterility, dwarfness, etc.)
were removed from the plots during the X z to X 4 generations. 75 plantsfrom each plot were taken at random from the X 4 , and the resulting X 5lines were tested in an experiment of split plot design with three replications. Heading date and plant height were observed.
The results of this experiment showed that (1) the distribution wascompletely normal in all plots, (2) the control and X-rayed plots had thesame population mean, and (3) the standard deviation became larger due
to irradiation. Further, selection in both directions from the X s populations showed that X-rayed plots had larger heritability values than thecontrol plot. These facts may indicate that in polygenes for quantitative* Ohara Institute for Agricultural Biology.

96

RESEARCHES CARRIED OUT IN 1957

characters, mutations with plus and minus phenotypic effects occur equally frequently. The rate of increase of genetic variance due to 1,000 r
irradiation was 0.015 (day) in heading date, and 0.084 (cm) in plant height.
(c.f. Jour. Heredity, 49: 11-14)

65. Effects of r-radiations upon Germination and Growth of Rice Plants(By T. OHTA, S. KAWASHIMA and S. MATSUMURA)

Dormant seeds of paddy rice were exposed to r-rays by 60CO at 5-70 kr.
Just after irradiation they were sown in an air-conditioned greenhouse at
28°C and 80% humidity. There was no marked difference in the germination rate between untreated and treated seeds at less than 40 kr, but thegermination rate was reduced to 60% at 50 kr and further to 22% at 70
kr. The number of surviving seedlings a month after sowing decreased
with 'the increase of the dosage of more than 30 kr. It is assumed that
LD-50 and LD-I00 are about 45 kr and 70 kr, respectively. The growth ofseedlings from irradiated seeds was delayed in proportion to the dosageof more than 20 kr.
In some seeds stored about 2 months at room temperature (about
15°C) after irradiation, the effect of r-rays upon the germination andgrowth of seedlings seemed to be more pronounced at higher dosage (50kr or more). Also, when 30 day storage at O°C was used after irradiation,
the germination and growth of seedlings were delayed about 10 days,
compared with 25°C storage. When the seeds irradiated at 30, 50 and
70 kr were sown at 20°C and 28°C, the growth of seedlings at low temperature was more delayed and the effect of r-rays at 50 and 70 kr was
smaller than at high temperature. In seeds one year old LD-50 is between 50 and 70 kr and radiosensitivity may decrease with the aging ofseeds.
Without the soaking (water content, 13.7%) and after the soaking for
4 hrs (21.9%), 12 hrs (27.4%), 24 hrs (31.1%) and 48 hrs (35.0%), the seeds
were exposed to r-rays at 5-50 kr. Radiosensitivity in respect to thegrowth of seedlings increased with the increase of water content. It was
sudden in the seeds irradiated after 4 hrs soaking, increasing later gradually with the increase of soaking time.
When the seeds were soaked
in 0.75% solution of NaCI for 72 hrs and then irradiated, they could not
germinate at all even at 40 kr.
Young seed roots about 7 cm long were irradiated. At 4 kr the elongation of roots was inhibited and at 12 kr the lateral roots were nearly all
inhibited.

RESEARCHES CARRIED OUT IN 1957

66.

97

Radiosensitivity and Mutants in Barley Induced by Irradiation(By S. MATSUMURA and T. FUJII)

Dormant seeds of several commercial malt varieties of Hordeum distichum
were subjected to X- and r-ray treatments. X-rays were applied without
filter, at 160 KVP, 3 rnA and 10 kr, and r-rays were applied with 60CO, at
10, 20 and 60 kr. Radio-sensitivity of different varieties was compared as
to germination and survival rates and the growth of seedlings. Hordeum
had generally a lower sensitivity than Triticum. There was no differencein the germination rate between untreated and treated seeds (X -10, r-10 and r-20 kr). In two varieties with more sensitive germination,
Early Golden Melon and Asahi No.5, the germination rate was reduced
to about a half of that of untreated seeds at r-60 kr. On the other hand,
the growth of seedlings was slightly delayed at X -10 and r -10 kr, andwas markedly inhibited at r-60 kr, and most of the seedlings died before
heading in Asahi No.5. There was a marked difference concerning
survival rates between different varieties.
Various kinds of chlorophyll mutants, such as albina, chlorina, viridoalbina, etc., were found among the seedlings of the X2 -generation. The
frequency of head progenies with mutations generally increased in proportion to the radiation dosage. At the same dosage (10 kr), X-irradiation
was more effective in this respect than r-irradiation. Three varieties,
Early Golden Melon, Asahi No.5 and Spratt Archer, had the highest rateof chlorophyll mutants, while it was lowest in Extra Plumage and Plumage Archer with lower survival rates. It is assumed that the effect ofradiation on physiological and genetic conditions is different with different
kinds of radiations and varieties used.

67.

Effect of AET upon Radiosensitivity of Wheat Seeds(By S. MATSUMURA and S. KAWASHIMA)To examine the protection effect of AET (Amino-ethyl-isothiuronium)
upon radiosensitivity, seeds of Triticum monococcum flavescens were treatedwith 0.0001 and 0.001% AET-solution for 24 hrs just before X- and rirradiations at 5 and 9 kr.
Seeds treated with 0.001% solution showed no higher germination rates
and no better growth of seedlings than the water-soaked ones, while whentreated with 0.0001% solution rather better germination and growth ofseedlings were observed. X-rays were more effective than r-rays. At
X-irradiation, especially when 0.001% treatment was used, AET unexpec-

RESEARCHES CARRIED OUT IN 1957

98

tedly increased the inhibiting effect of radiation. In r-rays the protection
effect of AET was generally not found at 9 kr, while only the treatment
with 0.0001% AET at 5 kr slightly promoted germination and growth.
This promotion might be due to the effect of 0.0001% AET, mentioned
above. The protection effect of AET upon radiation could not, therefore,be easily determined.

68.

Diffusion of Radioisotopes in Soaked Wheat Seeds(By S. KONDO, N. YOKOTA and S. KAWASHIMA)

A simple method used to induce mutational changes in plants by radiations is to soak their seeds in water solutions containing radioactive isotopes. These solutions shall be called by their abbreviation; RI solutions.
This method is, however, accompanied by difficulties in the measurementof given doses.
The soaking experiments were performed in a room of constant temperature of 21±0.5°C with dry seeds of wheat, Triticum monococcum flavescens, soaked in RI solutions of pH 6 to 7 which contained 10/Jc/cc of 32p,
13 11, 905r, mcs, 65Zn, each.
Each set of experiments comprised 10 seeds
soaked in an RI solution of 2 cc. The changes in the radioactivity of theseeds were detected by a G-M
counter and X-ray film. The
number of counts of RI solutions
was measured by a Philips G-M
counter, Model 4035, using amica window G-M counter at
400'13.5 cm distance, with a work30J
ing voltage of 725 volts. After~200
soaking the seeds in an RI
solution for a given time, eachseed was rotated one revolution
water abJrrptwn
on a filter paper and thencounted as an intact seed. To
650eliminate the self-absorption
40
error in counting, each seed30
was ground into a fine powder
and then counted. The count
2
3 4
1/4
1/2
!
per minute (cpm) for theseTime of sool""9 Ln days
methods are denoted by A andFig. 1. Absorption curves for wheat seeds
B, respectively. The cpm's of
soaked in radioactive solutions.
A and B were compared with the cpm due to the RI contained in the

!oJ
500~

RESEARCHES CARRIED OUT IN 1957

99

outside solution, C. The amount of the outside solution was exactly
equal to the weight increase of the seed at that time. Then, BI A or
CIA can be used as a measure of the average concentration of the RI in
the water absorbed by the seed. The amounts of RI solutions diffusedin the wheat seeds per days soaked are shown in Fig. 1 and Table l.
The water absorption shown in Fig. 1 is indicated by the average weight
Table 1.

The concentration of 32p on, and inside the soaked wheat seeds,

T. monOBOBcum jlavescens, relative to the outside solution.

Surface and inside radioactivityDayssoakedOutside radioactivity, A

1/4
1/2
1
2
3
4

500 cpm/seed
797
"
1030
"
1420
1750
"
1820

"

Beforegrinding, B

Aftergrinding, C

99cpm/seed
120
"
147
"
384
"
483
"
387

145 cpm/seed
164
"
171
"
452
"
601
"
505

- -

"

--

"

Relative activity
B/A

C/A

0.20
0.15
0.14

0.29
0.21
0.17

0.27
0.28
0.21

0.32
0.34
0.28

The symbol A denotes the cpm/seed of the outside solution of an amount equal
to the increase in weight of a soaked seed; Band C those of wheat seeds after
soaking as an intact seed and then being ground, respectively.

Fig. 2. Macro·autoradiographs of wheat seeds soaked 3 days in 32p and1311 solutions.

RESEARCHES CARRIED OUT IN 1957

100

increase per seed for 50 seeds. From Table 1, we find that the RI concentration inside the seeds in lower than in the outside original solution
and that the quick measurement of cpm of intact seeds results in a
relatively good approximation to the true cpm of ground seeds. In fact,
the macro-autoradiographs in Fig. 2 show that RI solutions are mostly
Table 2.

A comparison of adsorbance ratios of radioisotopes on the soaked
wheat seeds, T. m07l1Jcoccum flavescens.
90S r

Days soaked

1/4
1/2

8.06

1

4.47
3.14
2.97
2.57
3.24*

2

3
4

4*

1.33
4.33

0.83
0.83
0.78
0.70
1.83*

0.20
0.15
0.14
0.27
0.28
0.21
0.28*

i.
All the values are expressed in B/A except for those with asterisks which are
in CIA. The B/A and CIA are defined in Table 1.

adsorbed on the seed coats and that the RI concentrations inside theseeds are very low for 1311 and 32P. The abrupt change in the absorbed
amount of 32p shown in Fig. 1 may indicate that the biological functionof the seed coats changes between one and two days after being soaked.
This change could not be detected by 1311. An interesting fact is shownin Table 2 that 65Z n , mcs and 90S r are very stnmgly adsorbed on wheat
seeds compared with 32P.
We may thus conclude that the seed coats of wheat are serving asfilter for radioisotopes. It should be noted that the uptake of RI solution
by plants may always be performed only through such semipermeable
membranes. If such membranes could be synthesized, they might be used
for decontamination of RI contaminated solutions. Due to the semipermeability of the seed coats, the dosimetry of soaked seeds in 32p or 1311
solutions can be made with about 10% accuracy only if the radiations fromoutside the seeds are taken into account. In the case of 65Z n, 90S r , andmcs, the adsorbed or absorbed amounts also considerably contribute to
the dosage given to seeds.

RESEARCHES CARRIED OUT IN 1957

K.
69.

101

GENETICS OF MICROORGANISMS

Transductional Analysis of Alonophasic Types of Salmonella(By Tetuo IINO and Joshua LEDERBERG*)

The specificity of flagellar (H) antigen in Salmonella is controlled by
two distinct loci, phase-l by H 1 and phase-2 by H 2 • Which one is manifested in a given clone depends on the phase determinant at the H 2 locus.
That is, the alternation of H 2 state leads to the alternative expression,
which has been known as phase variation of H antigen (the Annual
Report, 1956).
Some Salmonella strains do not express two phases but only one. Thosestrains· are called monophasic-l or -2 strains depending on their fixedphase, either phase-l or phase-2 respectively. Three additional groups ofthe genes which are involved in the production of H antigen were discovered by transductional analysis of the monophasic strains.
S. abortus-equi CDC-26 is stable in phase-2, enx-type. A rare alternative
phase can occasionally be obtained by antiserum selection, resulting in an
equally stable phase-I, a-type. Transductions were carried out from enxphase of CDC-26 to i-phase of a diphasic strain of S. typhimurium, TM2
i : 1.2 (such transduction is designated CDC-26 (a) : enx-x TM2 i : 1.2).
Among 65 transductional clones which had been selected on semisoli
nutrient gelatine-agar media (NGA), 4 expressed diphasic a : 1.2 type, 42diphasic i: enx and 19 monophasic-2 enx which carried a hidden H 1 '.
Thus, when a is transduced the resultin transductions remain diphasic,
whereas when enx is transduced a fraction of the transductional clones
becomes monophasic. By anti-enx selection, i-phase cultures were obtained
from the monorphasic enx-transductional clones. The i-phase cultures
obtained were also monophasic. The stabilization of the H 2 state in S.
abortus-equi is therefore caused by a factor which is linked to H 2 • The
factor will be given a symbol Vh 2 -.
S. typhimurium SWI061 is a monophasic-2 mutant of a diphasic strain
TM2 i: 1.2. The culture reacts to anti-1.2 serum but not to anti-i serum.
However, the strain frequently produces nonmotile H-negative (non-flagellar) subclones, which in turn revert to motile cells with 1.2 antigen insuccessive cultures. From the transduction, diphasic S. abony CDC-I03
b : enx-x SWI061, monophasic-2 enx, diphasic b : 1.2 and a a small number of i : 1.2 types were obtained. The change from the monophasic type todiphasic types was always coupled with the loss of the ability to oscillate* Department of Genetics, University of Wisconsin, Madison, Wis., U. S. A.

RESEARCHES CARRIED OUT IN 1957

102

between motile and non-motile types. These results are consistent withthe following explanation. In SWI061, HI i is inactive; on the other handH 2 1.2 changes its state as in usual diphasic strains. Consequently, whenH 2 is active, phase-2 antigen, 1.2, is produced, and when H 2 changes toinactive, that is both HI and H 2 are inactive, the cell produces H antigen
and becomes non-motile. The production of diphasic i : 1.2 type suggeststhat the inactivation of HI is not caused by an intrinsic change' of HI
itself but by an inhibition of its function by a gene linked to HI. The
linkage and the recombination between HI and the controller of HI-activity
were confirmed on Flal + (linked to HI) transduction SWI061-x S. heidelberg
FlaI- r : 1.2, from which monophasic-2, -(i): 1.2, and diphasic i: 1.2 aswell as diphasic r : 1.2 were obtained. The controller of Hractivity isdesignated Ah i -. Ah I- was discovered in two other monophasic-2 strains ofS. typhimurium, SW629 an SW547. All of three Ah I- are non-allelic andlinked to HI and Flal. The parallel type of gene, Ah 2 - , which inhibits
the function of H 2 was discovered on four monophasic-l strains of S.
typhimurium. All of them are linked to H 2 • Both Ah l and Ah 2 are phase
specific but not concerned with the specificity of antigen types, which aredetemined exclusively by HI and H 2 •L.

MATHEMATICAL GENETICS

70.

Chance of Fixation of Mutant Genes(By Motoo KIMURA)

Because of its importance in evolutionary genetics, the probability offixation of mutant genes has been studied by Fisher, Haldane and Wright.
However, due to mathematical difficulties involved, so far only a few
cases have been successfully worked out.
In the present report I will present a quite general solution in which
an arbitrary degree of dominance is taken into account.
Let N be the effective size of a population and let sand sh be theselective advantages of the mutant homozygote AA and heterozygote AA'
respectively over the wild type homozygote A'A', then the probability offixation of a mutant gene A which has appeared in the population may
be given by

where c=Ns and D=2h-1. For an advantageous but nearly recessive
mutant gene (s>O, O<h<l), the above formula may be substituted by

103

RESEARCHES CARRIED OUT IN 1957

u=e-2NS"2/(1-2")/2Y(~~2hl / {1-2(/)(v4Nsh2/(1-2h))},unless 2Ns is small.

Here (/)(x) stands for the error function(/)(x) = (I/V21r)): e-

z2 2
/

dx.

As an example, consider a case with N= 10 3 and s= 10- 1 • If the mutant
gene is completely recessive (h=O), u=0.8xlO- 2. With slight phenotypic
effect of h=O.01 in the heterozygote u=0.9xl0- 2 , while with h=O.I,
u=2.3xlO- 2.
(d. KIMURA, M. 1957. Ann. Math. Stat. 28:882-901)

71.

Effect of Natural Selection in a Finite Population(By Motoo KIMURA)

In the last report of our institute, I have presented the results of investigation on the interaction between natural selection and random genetic
drift, assuming complete dominance between a pair of alleles. The present
report concerns with a more general case with an arbitrary degree ofdominance.
Consider a pair of alleles A and AI with respective frequencies of x
and l·x in a population of effective size N. If we designate by s, sh ando the fitnesses of three genotypes AA, AAI and AIAI respectively measured in Malthusian parameters, then the probability density, ifJ(x, t), thatthe frequency of A lies between x and x+dx satisfies the following partial
differential equation:
aifJ =~~Lxil--'~ifJ}_-~{Sx(l-x)[h+(1-2h)x]ifJ}.
at
ax2 I 4Nax

The smallest eigenvalue Ao giving the rate of steady decay may be expanded
into a power series in Ns as follows:
2NAo= 1+ K 1 (Ns)+ K 2(Ns)2+ K g(Ns)3+ K 4(Ns)4+ .... ,
where
1
1
22 .3.,
K 1 = -5 D , K 2 = 2.5+ 53.7 D",

122
1 - - - __-D2
73
K =---D-----D3
K 1= - - 3

2.53.755.7'-

23.53.7

2.33.55

22 .35 D4
56 .7 3.11'

etc, in which D represents degree of dominance such that D=2h-1. Thus
for the case of no dominance D= 0 and for the case of complete dominance
D= 1 or -1 according as A is either dominant or recessive.

104

RESEARCHES CARRIED OUT IN 1957

72.

Zygotic frequencies in a Partially Self-fertilizing Population(By Motoo KIMURA)

In higher plants, practice of self-fertilization is quite wide-spread, withthe result more homozygotes may be found in a population than would
be expected under exclusive random mating.
We shall consider an infinitely large population and designate by R theprobability that an ovule is fertilized by a pollen taken as a random
sample from the entire population and by S( = 1- R) the probability that
it is fertilized by a pollen from the same plant. Thus Rand S may betermed the proportion of cross- and self-fertilization respectively.
(i) One locus segregating: Let AI, A 2,
be an arbitray number ofalleles with respective frequencies of Xl, X2, ..
Assuming that mutation,
selection and migration are absent, the equilibrium frequency of thehomozygote AiA i may be given by
F {AiAi}=}Rxi2±SX~
r
2R+S '

while the frequency of the heterozygote AiA j (i~j), may be given by
4RxiXj
Fr{AiAj}=ZR-tS'
If the population is not in equilibrium, deviations of zygotic frequencies
from their respective equilibrium values may be decreased at the rate of(S+2R)/2

per generation.
(ii) Two loci segregating: Let Xi be the frequency of the i-th allele Ai in
the first locus and let yk be that of the k-th allele B k in the second locus.
If r is the recombination fraction between these two loci, the equilibrium
frequencies of double homozygote AiAiBkBk, single heterozygotes AiAiBkBl,
AiAjBkBk ane double heterozygote AiAjBkB l are given respectively by
F {A AB B }=2RxI2Yk2+SXiYk
r i , k k
2R+S

2RSLxlYk(1-xi)(1-Yk)(2R+S)(2R+S+LS)F {A AB B}= 4RxlYkYI[(21?+S)xi+LS1_ (k~l)
r ilk 1
(2R+S)(2R+S+LS)F {A A B B }=_41?:l7iXjYd(2R+S)Yd-/ R
r i j k k
(2R+S)(2R+S+LS)J

andF {A -A B B}= 8Rx/..Xj}jkYl (i~J', k~l),r, } k I
2R + S + LS

(i~')
J

RESEARCHES CARRIED OUT IN 1957105

where L=2r(l-r). These results show that distributions of zygotic frequencies between two loci are not independent under partial self-fertilizationeven when the two loci are on different chromosomes. For example, with
two unlinked loci each containing a pair of alleles in equal frequencies
the coefficient of departure from random combination of homozygous
phases, i.e.

O(ii'kk)=f!"J/liAiIi~13k} _ =1+!?(l=RJ__

Fr{AiAi}Fr{BkBk}
3+R
is always larger than unity, giving the maximum value of 1.07 when
R= 2V3-3=0.464.

73.

Overdominant Genes in an Partially 5elf-fertilizing Population(By Motoo KIMURA)

Behavior of heterotic genes in a random mating population has beenrather well understood, but the one in a non-random mating population
has not yet been fully analysed. In plants, however, self-fertilizing
mechanism is quite wide-spread and investigation into the behavior ofheterotic or overdominant genes in a partially self-fertilizing population
should be required.
Let R be the probability that an ovule is fertilized by a pollen taken as
a random sample from the entire population (assumed to be very large)
and 5 (= 1- R) be the probability that it is fertilized by a pollen produced
from the same plant. Consider a pair of alleles Al and A 2 and designate
the selective values of three genotypes AlA], A l A 2 and A 2 A 2 by l-s 1 , 1
and l-s 2 respectively, then one of the following conditions must be satisfied in order that the alleles Al and A 2 are maintained in stable equilibrium:( i) If Sl, s2>1/2, a stable equilibrium is possible for any degree of selffertilization.
(ii) If Sl =S2= 1/2, the proportion of self-fertilization must be smaller
than 1, i.e. 5<1.
(iii) If at least one of Sl and S2 is less than 1/2, the proportion of selffertilization must be restricted such thatassuming here that Sl > S2.
If one of the above conditions is satisfied, the equilibrium genotypic
frequencies may be given by

RESEARCHES CARRIED OUT IN 1957

106

Pu=(l- j)X1 2 + j X1
2P12 =(1- ])2X1X2
P 22 =(1- ])X22+ jx, ,1where Pu, 2P12 and P 2, are the frequencies of A 1A 1, A 1A 2 and A,A 2
respectively and Xl and £0 2 are the equilibrium frequencies of A 1 and A 2such that

X1=_i s_ v (l-+
1- 1

Sl

f),

x2=1- x 1,

S2

in which i.=SlS2/(Sl +S2) andj=4~ [2(1-i.)-(1-2i.)S- v Dl ,
z.

where

D=4(1-i.)2+(1-2i.)2S'-4(1 +i.)(1-2i.)S .

It should be noted here that j is different from the ordinary inbreeding
coefficient defined for neutral genes which isS
1=2R+S
under partial self-fertilization. For example, with S=R=0.5, Sl=S2=0.5,
the condition (ii) is satisfied and j = (5- 17)/4=0.22 which is different
from 1 = 1/3=0.33.

v

74.

On the Decrease 01 Fitness due to Deviation Irom Optimumin Quantitative Characters.
(By Motoo KIMURA)

It has been confermed for several simple quantitative or meteric characters such as height and weight that phenotypically extreme individuals(deviants) are less fit than those near the population average.
The present report is an attempt to treat this problem based on the assumption that for each locus concerned to a metric character, fitness ofeach gene combination is determined independently of its contribution to
the metric character.
Consider a locus with an arbitrary number of alleles, say, A 1 , A 2 , ••• An
and designate the frequency of homozygote AlAI by Pi! and that ofheterozygote AIA j (i~j) by 2Plj. We assume that a large number of

RESEARCHES CARRIED OUT IN 1957

107

loci are concerned for the determination of the metric character. If Y is
the measure of the metric character which distributes normally with mean
Y and standard deviation 11 then the average fitness of individuals withmeasurement Y may be given by__ (Y_Y)(Y_Y)2_t1 2a(Y)=a+ ~-22: l1ay+
211 4
2: f1.12(a, y) ,(1)

where a is the average fitness of the population and the summation isover all relevant loci. In the above formula l1ay and f1.12(a, y) are given by
l1ay= 2: alj(Yij-y)Pij
ij

andf1.12(a, y)= 2: (aij-a)(Yij-y)2Pij
ij

in which aij and Yij stand for the contributions of AiA j to the fitness andthe metric character respectively.
If 2: f1.da, y)<O, there exists an optimum value of Y for which a(Y)takes its maximum value. Thus we obtaina(Y)-a(Yopt ) =(Y -I1YoPtY_l.. 11/ ()
2

11

2
'

where Yopt is the optimum value of Y, 11//112 is the heritability of metriccharacter and () is given by()= -2: f1.12(a, Y)/l1 g 2 .

As an important application of the above result, we assume that in eachlocus concerned a mutant gene is maintained in equilibrium due to counter·balancing effect of mutation and selection. This leads to(}=I/(I/lif,
where h is the selective disadvantage of the mutant heterozygote and(l/h) means the average value of l/h weighted according to the additive
genetic variance of the metric character in each locus.

M.
75.

TECHNICAL NOTE

Dose-rate Distribution in the 60CO Gamma Ray Room

(By S. KONDO and S. MATSUMURA)

The facility for

60CO

gamma ray irradiation was built in May, 19561 •108

RESEARCHES CARRIED OUT IN 1957

The plan of the gamma-ray room and the sectional view of the oOCo container are presented in Figs. 1 and 2.
The lead filters shownin Fig. 2 were designed
gamma room
to produce three regionsof roughly constant but
different dose rates. For
simplicity's sake in thedesign only the primaryradiation was taken into
account. The dose-rate
distribution on a horizontalline passing throughthe point 20 cm below thesource was measured witho -'----"'2ma solid CdS: Ag crystal
Fig.
1. Gamma ray irradiation room of the National
detector having 5 mm in
Institute of Genetics.
diameter and 23 mm inlength. Fig. 3 shows that the first and second filters give fairly goodplateau regions, constant to about ±5% for annuli at least 5 cm wide atthe distance 20 cm below the source, whereas the third filter provides noreal plateau. Apparently when the attenuation factor is greater than 10,
secondary radiations and boundary effects of the filter must be taken into
account in the design of such afacility.
a 5
The total emission of the source
was determined in two ways.
With the source 200 cm above
the floor an inverse square curv,ewas taken with the FRICKE ferrous
sulphate chemical dosimeter".
The dose rate was also measured
at a single distance with a
Victoreen ion chamber. The
effective source strength S incuries was calculated as:
Fig. 2. The sectional view of the container'-'of oOCo.

where d is the measured dose rate in r/hr at x cm from the source andone curie of 60CO is taken as producing 1.35.10 4 r/hr at 1 cm. The values
obtained are given in Table 1; they indicate good agreement between the

RESEARCHES CARRIED OUT IN 1957

109

two methods. Our
best estimateofr/hr
the effective source r/hr/00/000
strength was 31.3
curies as of Nov.
1, 1957. For thesemeasurements
theampoules containing
500
50
the ferrous solution
were covered by a
the magnified
3 mm thick lead petri
curveddish to eliminate
secondary and back2
Omscattered radiations
Horizontal distance from 60CO source in cm.
from the 60CO container. Table 1 also Fig. 3. The distribution curve of dose rate along a
horizontal line 20 cm below the 60CO source.
shows that the inThe measurements were made with the use of averse square law
small CdS gamma meter on December 24, 1956.
holds well from 9 to
60 cm from the source. If, however, the ferrous dosimeter is unshielded, the apparent source strength varies appreciably when the detector
is closer than 20 cm to the source or farther than 160 cm from it.
Table 1.

Measurements of the effective source strength by FRICKE ferrous
dosimeter and Victoreen ion chamber.
Effective source strength in curies

Distance between
center of detector
and center ofsource9.2cm
11.6 "
18.2 "
31.2 "
60.0 If

FRICKE ferrous sulphate
Direct result

Corrected forlead shielding

26.9
26.5

31.0
30.6
31.4

27.4
27.6

Victoreen ion
chamber

31.8
31.2

The degree of hazard to operating personnel was checked by the following measurements. When the source was at the bottom of its 100 cm deepstorage pit, the radiation at the floor level above the pit was initially
170 mr/hr which was reduced to 13 mr/hr by the addition of a 3 mm thick

110

RESEARCHES CARRIED OUT IN 1957

shield plate. The average dose rate from scattered radiations at theentrance, when there were no doors, was 4 mr/hr.
1) MATSUMURA, S.: 1956. Radioisotopes 5: 60-64.
2) WEISS, J., A. O.
ALLEN and H. A. SCHWARZ: 1956. Proc. Peaceful Uses of A. E. 14, 179. 3)
National Bureaus of Standards, U. S., Handbook 54, p. 42 (Sept. 1, 1954).

76.

Temperature Dependence of the Window Glass Dosimeter1(By Sohei KONDO)

Measurement of gamma ray dose by means of changes in the absorption
coefficient of ordinary window glass has been reported previously2. To
improve the accuracy of this glass dosimetry method, the temperaturedependence of the glass dosimeters has been empirically investigated. In
general irradiated glass loses its induced color at a rate strongly dependent
on the storage temperature. The following equation of fading obtains forwindow glass irradiated with 60CO gamma rays at 22°C
D(t, -r)=D(O, -r)f(t/-r) ,(1)

where D(t, -r) denotes the value of D(O, -r) at t hours after the end of irradiation, D(O, -r) being the increase in absorption density immediately
after the -r-hour irradiation*. The fading factor f(t/-r) can be expressedby:3.
f(t/-r) =(l+t/-r)H"-(t/-r)l-A, (2)where the fading index ..l follows the relation:

3.0..l=exp(-11+0.03 T),

/5

250

300
Temperature ('K)

Fig. 1. Relationshipbetweenfading index ,\ given in (2) andtemperature T.

(3)

and T is the temperature in degrees
Kelvin.
The experimental data (Table 1 and Fig.
1) show satisfactory agreement withequations (2) and (3). The relation between
the total dose R given to the glass at 22°C
and the increase in absorption density
D(O, -r) of the glass is given by:
R= (1222±2)D(O, -r)-r 0 • 154 ,(4)* Absorption density D is defined by 1j1o=10-DX where I and 10 are the intensities of transmitted and incident radiations, for the present case with wave length
4200 A; and x denotes the thickness of the glass dosimeter.

111

RESEARCHES CARRIED OUT IN 1957

where R, D(O, ,,) and" areexpressed in kr, cm- 1 andhours, respectively. The
system of glass dosimetry
employed using equations(1) to (4) yields an accuracyof ±2%.
Fig. 2 shows the comparison of the response of
the glass dosimeter irradiated at 22°C with thatat 89°C. From this experiment we conclude thatthe fading index A in equation (2) differs from thepower index on " in e100
. 50
quation (4). The latter!l:lse R
causes the non-linearity in
Fig.
2.
Dependenceofabsorption
density increase
the relationship between Rof glass on irradiation temperature.
and the induced absorption
increase D even well below the saturation of D.
Table 1.

Fading of relative absorption coefficient, D(t, r)jD(O, T), with time
t for T=16hr at 22°C with dose rate about 3 krjhr.

~~._ t (hr)°C~Fading~

1

4
( +5)

8.5
(+10.5)

23
(+25)

46
( +48)

0.926+
0.930+

0.891+
0.909+

0.838+
0.848+

0.795+
0.785+

0.880
0.854

0.826
0.798

0.729
0.706

0.666
0.659

0.608
0.582

i 0.750

0.657
0.625

0.519
0.532

0.428
0.442

0.376
0.391

---------~~

16°C.1=0.1

Calculated
Ob::erved

0.973

36°C
.1=0.18

CalculatedObserved

0.947

58°C
.1=0.35

Calculated I 0.875
Observed I (**)

**

63
(+71)

(**)
(**)

I

0.732

-

~

0.766+
0.746+--- - - - - - - - - - - - - - -Adjusted to equal the observed values.

1) The main part of this report was published in J. Phys. Soc. Japan, 2: 224(1958). 2) S. KONDO: Ann. Rep. National Inst. Genetics, Japan No.7: 98(1957).

112

BOOKS AND PAPERS WRITTEN BY STAFF MEMBERS

BOOKS AND PAPERS PUBLISHED IN 1957
BY STAFF MEMBERS
ENDO, T. 1957. Column chromatography of anthocyanins.
FUJII, T. 1957. Studies on chlorophyll mutants in diploid wheat induced by radiation. W.I.S. No.6: 8.
------ 1957. Mutations in einkorn wheat induced by X-rays, III. Double recessive plants in a cross between basi-viridis II and chlorina. Seiken Zih6 No.
S: 33-36.
HORIKAWA, M. 1957. Growth, differentiation, and tryptophan metabolism in eye
discs of D. melanogaster in tissue culture. D.I.S., 31: 124.
KAWAHARA, T. 1958. Studies on heterosis in the domestic fowl, I. Viability andresistance to blackhead in the F hybrid between two breeds of fowls. (In Jap.)
Jap. J. Zootech. Sci 28 (3) 181-184.
KIHARA H. (edited) 1957: Peoples of Nepal Himalaya. Scientific results of theJapanese expeditions to Nepal Himalaya 1952-1953 Vol. III, P. 425.
---- K. YAMASHITA, M. TANAKA and S. SAKAMOTO (1957): Geographical distribution of 4x and 6x forms of Aegilops crassa. 5: 11-12.
KIMURA, M. The mathematical theory of population genetics (In Jap). Iwanami, Tokyo.
--- The rate of change of ,population fitness by natural selection. Proc. International Genetics Symposia (Tokyo). pp. 467-471.
Some problems of stochastic processes in genetics. Ann. Math. Stat.
28: 882-901.
MATSUMURA, S. 1957. Induced mutation. (In Jap.) M. SHISA & N. KONDO ed.:Breeding of flowers and vegetables: 76-90. Tokyo.
(ed.) 1957. Equipments for control of temperature, humidity and light,
and their utilization. GihOdo, Tokyo. pp. 289.
- - 1957. Genetic and cytological studies on the genus Beta. VI. Comparative experiments with different triploid combinations, especially American
resistant varieties contribution the diploid parent. Seiken Zih6 No.8: 75-82- - - & T. FUJII. 1957. Genetical effects of ionizing radiation in einkorn
wheat. Proc. 1st Symp. Atomic Energy No.4: 691-696.
.- - - _ .
1957. Genetic effects of ionizing radiation in einkorn wheat.
W.I.S. No.6: 6-7.
- - - - - - - &. S. KONDO. 1957. Effects of X- and r-radiations upon wheat
seedling and their modification due to temperature or polyploidy. W.I.S. No.
6: 7-8.
- - - MOCHIZUKI, A. & S. SAKAMOTO. 1957. Triploid sugar beets and theirseed production. Proc. Intern. Genet. Symp., 1956: 314-317.
MORITA, T. 1957. Purine catabolism in D. melanogaster. D.I.S., 31: 137.
NAKAO, Y., Y. TAZIMA, T. YAMAMOTO 1956. Mutagenicity of sarkomycin. Nature
178: 1466.
NAWA S., T. TAIRA, & C. OSHIMA, 1957. Pterine dehydrogenase and its coenzyme
in D. melanogaster. Drosophila Information Service (D.I.S.), 31: 146.

.,

BOOKS AND PAPERS WRITTEN BY STAFF MEMBERS113- - - - - 1957. Nonenzymatic conversion of the yellow pigment
found in D. melawgaster. Ibid. 31: 146.
OGAWA, Y., G. MIKAMI 1957. Effects of Na-glucuronate on function of rabbit liver
after partial hepatectomy. (In Jap.) Reports on Glucuronic acid. 1: 167-169.
- - - & - - - 1957. Effects of Na-glucuronate on free amino acids in rabbit
serum after partial hepatectomy. (In Jap.) Ibid. 1: 170-172.
- - - & T. KAWAHARA. 1957. Serological determination of actin developmentin early chick embryo. (In Jap) Kagaku 27: (5) 253.
- - - Y. ABE & K. FUJIOKA. 1957. Promoting activity of kinetin on Yoshidasarcoma cell division. (In Jap.) Ibid. 27: (8) 414-415.
- - - K. FUJIOKA & Y. ABE. 1957. Growth, differentiation and regeneration I.
Effects of Na-glucuronate on division of Yoshida sarcoma cells. (In Jap.) Med.
and Biology. 44: (3) 169-172.
- - - & T. Ishihara, 1957. Ditto II. The meaning of frequency of dividing
cells in the regenerating liver of rat after partial hepatectomy. (In. Jap.) Ibid.
45: (1) 12-14.
- - - - - & - - - - 1957. Ditto III. Mitotic activity in the liver of rats bearing
sarcoma. (In Jap.) Ibid. 45: (3) 110-113.
----- Y. ABE & K. FUJIOKA, 1957. Effects of kinetin on division of Yoshidasarcoma cells. Nature. 180. (4593): 985-986.
OKA, H. I. & C. C. Fang. 1957. Classification of rice varieties by discriminantfunctions. (Phylogenetic differentiation of the cultivated rice, XIV.) Jap. Jour.
Breeding, 6 (4): 245-248. (In Jap. with Eng. resume).
-_. .._.. & K. M. Lin. 1957. Genic analysis of resistance to blast disease in rice
by biometrical genetic method). Jap Jour. Genetics, 32 (1): 20-27.
------- 1957. Complementary lethal genes in rice. Phylogenetic differentiationof cultivated rice, XV.) Jap. Jour. Genetics, 32 (3): 83-87.
---_ 1957. Tendencies of characters to associate in lines derived from hybrid
populations. (Phylogenetic differentiation of the cultivated rice, XVI.) Jap.
Jour. Breeding, 7 (1): 1-6. (In Jap. with Eng. resume).
- - - 1957. Sterility and vigor of lines derived from hybrids between distantvarieties of rice in crosses with parental varieties. (Phylogenetic differentiationof the cultivated rice, XVII.) Jap. Jour. Breeding, 7 (1): 7-11. (In Jap. withEng. resume).
- - - & Y. C. Ru. 1957. Studies of drought resistance in rice varieties. Jour.
Agr. Assoc. China, New Series 18: 7-17.
1957. Change of population genotype due to gametic and zygotic selections in hybrids between distantly related varieties of rice. Proc. Intern. Genetics
Symposia, 1956: 482-490.
OSHIMA, C. 1957. Genetical studies on DDT-resistance in populations of Drosophilamelawgaster. I. Jap. Contrib. to the Study of the Insecticide-Resistance
Problem (for WHO): 40-45.
- - - & T. HIROYOSHI. 1957. Genetic studies of resistance to DDT and Nicotinsulfate in Drosophila virilis.. Ibid. 54-59.
1957. Genetical studies on DDT-resistance in populations of Drosophila

114

BOOKS AND PAPERS WRITTEN BY STAFF MEMBERS

melanogaster. I.

Ibid. 40-45.

- - - - & T. HIROYOSHI 1957. Genetic studies of resistance to DDT and nicotinsulfate in Drosophila virilis. Ibid. 54-59.
1957. Studies on DDT-resistance of Drosophila melanogaster from theview point of population genetics. Proc. International Genetics Symposia,
1956 428-434.
SAKAI, K. 1957. Studies on competition in plant. VII. Effect on competition ofa varying number of competing and non-competing individuals. Jour. Genet.,
55 (2): 227-234.
------ & UTIYAMADA. H. 1957. Studies on competition in plants. VIII. Chromosome number, hybridity and competitive ability in Oryza sativa L. Jour.
Genetics, 55 (2): 235-240.
1957. Theoretical studies on plant breeding technique with special regard
to autogamous plants. Proc. Internat. Genet. Symp., Cytologia, Supp!. Vo!.,491-497.
- 1957. Theoretical studies on plant breeding technique. IV. The estimation of genetic correlation between traits and its use in forest-tree breeding.
Jap. Jour. Breed. 7 (2): 83-85.
- - - - - 1957. Theoretical studies on plant breeding technique. V. Coefficientof relationship in the breeding of autogamous plants. Jap. Jour. Breed. 7 (2):87-92.
SUGAHARA, T., & M. NAKAMURA. 1957. On the technical and clinical problems
concerning high voltage radiography. Journ. of Nippon Hoshasen Gijitsu Gakkai,
Supp!. No.2, 22-56.
TUTIKAWA, K., SUCAWARA, Y. and TANAKA, T. 1957. Genetic effects ofchronic r-irradiation in mice: A preliminary report (In Jap.) Proc. First Symposium on Atomic Energy: 706-711.
TAIRA, T., S. NAWA, & C. OSHIMA. 1957. On the pterine dehydrogenase in eyecolor mutants of D. melanogaster. Japan Contric. to the Study of the InsecticideResistance Problem (for WHO), 164.
--- ----- - - - - & - - - - 1957. Pterine dehydrogenase in eyecolor mutant of
D. molonogaster. D.I.S., 31: 164.
TATEOKA, T. 1957 Notes on Some Grasses, III. Bot. Mag. Tokyo 70: 8-12.
1957 Ditto IV. Jour. Jap. Bot. 32: 111-114.
- - - 1957 Ditto V. Bot. Mag. Tokyo 70: 115-117.
1957 Ditto VI. Ibid. 70: 119-125.
1957 Miscellaneous papers on grass phylogeny (8). On the subfamily
Andropogonoideae. (In Jap.) Jour. Jap. Bot. 32: 11-17.
- - - - - 1957 Ditto (9). On the groups having primitive features of external
morphology, especially, Micrairoideae, Anomochloideae, Oryzoideae, Olyroideae,
and Bambusoideae. (In Jap.) Ibid. 32: 42-49.
1957 Ditto (10). Proposition of a new phylogenetic system of Poaceae.
Ibid. 32: 275-287.
TSUJITA, M. 1956. Genetical relationship between lethal lemon (1 eml) and lethal
albino in silkworm. Jap. J. Genet. 31 (12): 327-329.

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& C. MATUI. 1957 Lysogenic conversion of some genetic characters including sugar fermentation in Pseudomonas solanacearum. Suppl of Cytologia574-579.
K. WATANABE and S. TSUDA. 1957 Electron microscopy of thin sectioned
nuclei in Paramecium. Cytologia 22 (3-4): 322-327.
T AZIMA, Y. 1957. Genetic effect of radiation upon silkworm, 1. Effect of external
irradiation with f)-rays upon the germ-cells of the silkworm. (In Jap.) Proc.
First Symposium on Atomic Energy: 697-699.
- - 1957. Differences is sensitivity of germcells and chromosomes to radiationa nong some mutant strains of the silkworm. Proc. Intern. Genetics Symp.
1956 : 280-286.
1957. Use of radiation on animal breeding (In Jap.) Nogyo oyobi Engei
32 (4): 661-662.
YAMASHITA, K, H. KIHARA, 1. NISHIYAMA, S. MATSUMURA & K. MATSUMOTO (1957):
Polyploidy breeding in Japan. Proc. Int. Genet. Symp., 1956: 341-346.
YAMASHITA, K., H. KIHARA, 1. NISHIYAMA S. MATSUMURA, & K. MATSUMOTO. 1957.
Polyploidy breeding in Japan. Proc. Intern. Genet. Symp., 1956: 341-346.

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