THE XSSOCL~TION OF MUTANTS M'ITH HOMOZYGOUS
         DEFICIESCIES IN ZEA MAYS

                 R.\KH;\K;\ ~IcCLISTOCK
           ~-ni;`PrSity of MissntLri, Coimbia, Missowi

                 Received 1\Iay 3, rgqr

                        INTRODUCTION
T HE evidence to be presented in this paper supports the supposition
   that some recessive mutants in maize are caused by homozygous
minute deficiencies. A method by which homozygous minute deficiencies
may be produced in maize has been presented in a previous publication
(MCCLINTOCS 1938). It is related to the aberrant mitotic behavior of
ring-shaped chromosomes. It has been shown both in maize and Nicotiana
(Srrxo 1930; R. E. CLAUSES unpublished) that a chromosome in the form
of a ring does not maintain itself unaltered through successive nuclear cy-
cles. The details of this behavior have been presented in the two mentioned
papers. They may be summarized briefly as follows: (I) Ring-shaped
chromosomes may decrease in size by loss of segments of chromatin from
the ring. (2) Ring chromosomes may increase in size by duplications of seg-
ments of chromatin composing the ring. (3) Ring chromosomes may be
completely eliminated from sister telophase nuclei during a mitotic cycle.
(4) The frequency of these occurrences depends upon the length of the
chromonema composing the ring- the larger the ring chromosome, the
more frequent the aberrant mitotic behavior. In maize, aberrant mitotic
configurations producing alterations or elimination of the ring chromo-
somes may occur in 17 to 20 percent of all the division figures if the chro-
monema composing the ring is as long as that of the longest chromosome of
the complement. Ii the ring chromosome is small&for example, composed
of only four chromomeres~~the aberrant mitotic contigurations are very
infrequent and may occur only once in every five or six hundred divisions.
In all other nuclear divisions the ring chromosome behavior is normal;
the two sister halves of the ring chromosome separate freely and pass to
opposite poles of the spindle tigure alon g with the rod chromosomes of the
complement.
1J'hen aberrant configurations are produced by large ring-shaped chro-
mosomes, the chromatin content of the ring chromosome usually is altered
in each resulting sister telophase nucleus. Only rarely is the ring chromo-
some lost to one or both sister telophase nuclei. In contrast, aberrant mi-
totic contigurations produced by small ring-shaped chromosomes usually
result in total loss of the ring-shaped chromosome from both sister telo-
phase nuclei. Only rarely do the telophase nuclei receive altered ring-
shaped chromosomes.

GENETKS 25: j-12 September rglo


          HOMOZYGOUS DEFICIENCIES IX XI.\IZI<           543
The method by which ring-shaped chromosomes are lost to nuclei or

become altered has been described previously (MCCLINTOCK 1938). It may
be outlined briefly as follows. At some mitotic prophases the two sister
halves of a divided ring chromosome form a continuous double-sized ring
chromosome instead of two freely separating ring chromosomes (Prophase,
fig. I). It is not known whether this results from the method of reduplica-
tion of the chromonema composing the ring chromosome or from a somatic
crossover between the two sister chromatids subsequent to this reduplica-

Late anaphase

T&phase

Prophase

Late anaphase

Telophase

    7 2
Q
-e-----

T&phase

I      3

I 0 E

I I   I

DESCRIPTIONS OF TEXT FIGURES

FIGURFL I.-Diagram illustrating a method by which a ring chromosome becomes altered in
chromatin constitution. Upper left: a ring chromosome in a resting nucleus. The clear oval repre-
sents the centromere. The individual parts of the ring chromosome are designated by the numerals.
Upper middle: Aprophase configuration following a "crossover" between the txvo sister chromatids
of the divided ring chromosome. A dicentric, double-sized ring chromosome is produced. Upper
right: Appearance of the dicentric ring chromosome in the following anaphase. Breakage of the
chromatin strands between the centromeres may occur at any position. Three possible positions,
a, b, and c, respectively, are indicated by the dash lines, The resulting broken strands at late
anaphase and the new ring chromosomes formed at telophase by fusions of broken ends of these
strands are diagrammed below in the bracketed figures for the breaks a, b, and c, respectiwly.


544                  II.\KH.\K.\ ~I~~(`LIXTOCK
tion. At early anaphase. the two centromeres which are present in this
double-sized ring chromosome move toward opposite poles of the spindle
figure (Anaphase, fig. I). The subsequent behavior in the spindle figure de-
pends on the size of the ring chromosome. `The ring chromosomes used in the
investigations to be described in this paper were small. Therefore, the follow-
ing description will be confined to their behavior. !Vith such small double-
sized ring chromosomes, continued movement of the centromeres is US-
ally suspended and the double-sized ring chromosome remains in the region
of the equatorial plate during anaphase. It is consequently eliminated from
both sister telophase nuclei. Very occasionally, however, the strands of
chromatin between the two centromeres become broken, and a segment of
the double-sized ring chromosome enters each telophase nucleus. Fusion
occurs between the broken ends of the segment. Consequently, a newly
organized ring-shaped chromosome is produced in each of these telophase
nuclei. Illustrations of several types of ring chromosomes which may be
produced following breakage of a double-sized, two-centromere ring
chromosome are given in figure I. It will be seen that ring chromosomes
with duplicated segments or ring chromosomes with deticient segments
may be produced by this process. In either case, the size of the segment
may be minute or relatively large. Although aberrant mitotic configura-
tions, which could result in loss or change in size of the ring-shaped chromo-
somes, are relatively infrequent, it has been found that the frequency of
loss of the ring-shaped chromosome from both telophase nuclei is consid-
erably greater than the frequency of breakage of the ring chromosome with
inclusion of broken segments in each of the sister telophase nuclei. This re-
lationship is of considerable importance in the study to be described.
-1 simple method of obtaining sectors of a plant or even whole plants
which are homozygous deficient for a small segment of a chromosome may
be outlined as follows. It is necessary to have a rod-shaped chromosome
from which a relatively short segment has been deleted and a ring-shaped
chromosome composed of the chromatin deleted from the rod-shaped
chromosome. `The deficient rod chromosome plus the compensating ring-
shaped chromosome are thus equivalent in chromatin content to a single
normal rod chromosome. (See Df-1 and R-I; Df-a and R-z, tig. 2.) Cnion
of two gametes, each with a complete genomic complement but possessing
the deficient rod chromosome and its compensating ring chromosome (in-
stead of the normal rod chromosome), could give rise to plants with two
homologous deficient rod chromosomes and two homologous compensating
ring chromosomes. However, because of their small size, the ring chromo-
somes would irequently be lost to telophase nuclei following the formation
of aberrant mitotic configurations. Thus. in the development of such
plants, some nuclei would contain only a single ring chromosome following


           HOMOZYGOUS DEFICIENCIES IN MAIZE          54.5
loss of one of the ring chromosomes. During subsequent multiplication of
such cells, an aberrant configuration could eliminate the second ring chro-
mosome. Thus, cells would be produced whose nuclei have no chromatin
segment covering the deficiency in the rod chromsomes. These cells would
be homozygous deficient for the full extent of the deficiency in the rod
chromosomes. (For this preliminary presentation, it may be assumed that
these cells are inviable or incapable of further multiplication.) Very occa-
sionally, however, an aberrant mitotic configuration of a ring chromosome
would result not in loss but in a changed composition of the ring chromo-
some as illustrated in figure I. If, during development of the sporophytic
tissues, one of the ring chromosomes becomes reduced in size by loss of a
minute segment and subsequently, the second ring chromosome is lost to a
cell at a mitotis: anaphase, all the cells arising from this latter cell could be
homozygous deficient for a minute segment-the segment which had previ-
ously been deleted from the remaining ring chromosome. When the loss of
a minute amount of genie substance results in a change in the genotype
which is neither cell lethal nor interferes greatly with the capacity of such
cells to multiply, a sector of tissue could be formed with a visibly modified
phenotype. Minute deletions of different segments of the ring chromosome
could be produced following such aberrant mitoses. In one plant an aber-
rant mitosis might result in the deletion of one particular segment from the
ring chromosome. In another plant, an aberrant mitosis might result in
the deletion of an entirely different segment. Thus, tissues homozygous
deficient for different segments within the limits of the deficiency in the
rod-chromosomes could be produced. If, when homozygous, each minute
deficiency results in a particular type of visible modification of the tissue,
a number of different and distinguishable types of mutant sectorials
should be produced in these plants, each of which should be associated
with loss of a particular segment from the ring. Furthermore, sectors show-
ing identical characters should arise independently in a number of different
plants if sufficient numbers of plants are available for observation. This is
because a double-sized dicentric ring chromosome (fig. I) could be broken
at the same position on a number of independent occasions resulting in the
production of altered ring chromosomes from which the same segment has
been deleted.
If two adjacent segments, each of which when homozygous deficient
produces a mutant character, are simultaneously deleted from the ring
chromosome during an aberrant mitosis, a compound mutant sectorial
should be formed showing the characters caused by each deficiency-that
is, comparable to a condition of homozygosity for two recessive mutants.
If a mutant sector in a plant is included in the inflorescence? gametes could
be formed containing the deficient rod chromosome plus the deficient ring


546                  BARD.4R.i McCLINTOCK

chromosome. Fusion of two such gametes could give rise to plants which
are homozygous for the minute deficiency and thus homozygous for the
phenotypic expression which it induces. In such a way, altered ring chro-
mosomes may be isolated.

The appeurauce of the tissues of plants k&h two de,ikient rod chromosomes and two compensatiug
ring chromosomes, Ring I and Ring 2, colwnns one and tzo, respectively. S represents a ring chromo-
some whose chromatin completely covers the dejiciency in the rod chromosomes. a, b, and c represent
ring chromosomes with minute dejLiencies; eech defitiency, when homo;ygozrs, gives rise to the
phenotypic Lharacter a, b, and c, resbectively. a b and b c represent ring chromosomes with tu#o
swh minute dejiciencie., ;qhich, whelc home:ygous, give rise to the compowd mutant a b and b c,
respectively. The appearance of the tissues when both rings are present is shown in column 3. The +
indicates normal, non-m&ant tissues. In column5 4 and j the phenotypic appearance of the tissurs
following somatic loss of Ring I and Ring 2, respectively, is indicated.

RING I         RING 2

APPE.4RANCE OF   APPEARANCE OF   APPEARANCE Ok-
TISSUES WHEN   TISSUES FOLLOU--  TISSUES FOLLO\!
BOTH RIiVGS ARE    ING LOSS OF      INC LOSS OF
  PRESIXT        RING I         RIKG 2

ix
N
N
N
N
N
a
a
a
a
b
b
b
C
c
ab

K
3
b
C
ab
bc
b

a b
b c
c
ab
bc
llC
bc
bc

+
+
+
+
+
+
+
+
;
+
b
b
C

b

+
a
b

ab
bc
b
C
ab
bc
C
ab
bc
RC
bc
bc

+
+
+
+
+
+
a
a
a
t
b
b

C
ab

The meaning of simple and compound mutants produced by altered
ring chromosomes is important for an understanding of the logic of this
paper. Therefore, a table has been prepared (table I) to illustrate this
meaning. Assume three adjacent segments in the ring chromosome each
of which when homozygous deficient results in the characters a, b, and C,
respectively. If, in individual plants, altered ring chromosomes are iso-
lated with the simple deficiency mutants a, b, and C, respectively, and the
compound mutants a b and b C, respectively, combinations of each of
these altered rings with a normal ring chromc.some or with any other al-
tered ring chromosome may be made and should give predictable results.
These are shown in the table along with the characters of the tissues which
would be formed following mitotic loss of one or the other of the ring chro-
mosomes, respectively.


             HOMOZYGOUS DEFICIEXCIES IN M;\IZE           547
It is the purpose of this paper to describe the types of simple and com-
pound mutants which arise in plants possessing two homologous deficient
rod-shaped chromosomes and one or more compensating ring-shaped
chromosomes and to summarize the evidence which indicates that they are
caused by homozygous minute deficiencies.

T\f-0 CASES OF DEFICIENT ROD CHROMOSOMES PLUS COMPENSATING

RING CHROMOSOMES

Two cases of deficient rod chromosomes with compensating ring chromo-
somes were available for this study. (For a complete description of their
origin and behavior, see ~~CCILINTOCK 1938). In both cases, segments of the
short arm of chromosome 5 adjacent to the centromere were involved. The
two cases are illustrated in figure 2. In the diagram, the numbers I to 9 in
the normal chromosome j (first line in diagram) represent the positions of

                                           # .
Df-2                       0 , R-2
                                           *I*

FIGURE Z.-The upper figure is a diagram of a normal chromosome 5. The clear oval represents
the centromere. The region adjacent to the centromere in the short arm is designated by the
numerals I to 9. The middle figure represents a rod-shaped chromosome 5 deficient for the seg-
ments I to 4 (the Df-r rod-chromosome). The dotted line represents the extent of the deficiency.
To the right of this deficient rod chromosome, a ring chromosome is diagrammed possessing the
segment which has been deleted from the rod chromosome (R-I). In a similar manner, the Df-z
rod chromosome and its compensating ring chromosome have been diagrammed.

the first ninechromomeresadjacent to the centromere. The rod chromosome,
Df-1, is deficient for the four chromomeres adjacent to the centromere. Its
compensating ring chromosome (R-I) possesses these four chromomeres
(four chromomere ring below arrow, fig. E, Plate I). The Df-a rod chromo-
some (fig. 2) is deficient for these same four chromomeres plus the next five
chromomeres. The R-2 ring chromosome possesses these nine chromomeres
(fig. A, Plate I; lower arrow, fig. B and G I, Plate I). It should be noted
that the R-I ring chromosome does not cover the deficiency in the Df-2
chromosome but the R-2 ring chromosome does cover the deficiency in the
Df-1 chromosome. The transmission through gametes of the two deficient
rod chromosomes without compensating ring chromcsomes or with one or
more ring chromosomes is given in table 2.

The types of plants most used in this investigation and the types of tis-
sues which result from loss of the ring chromosome during development in
these plants are summarized in table 3. In all cases, the tissues are normal
when a single ring chromosome is present (either R-I or R-2) covering the
deficiency in the rod chromosomes. Tissues homozygous deficient for


548                 BARBARA McCLII\;TOCK

                          TABLE 2

Transmissions through gametes of Ike de/Gent rod chromosomes without ring chromosomes or with
  various ring rhromosomes. + represents transmission; - represents non-transmission.

THROCGH 0        THROUGH 3

   Df-r                         +                -
   Df-r plus R-I                    +             +*
   Df-r plus R-z                    +            +*
   Df-r plus R-I plus R-z               +             +*
    Df-z                              -                -
   Df-z plus R-r                         -                -
   Df-z plus R-z                    +             +
   Df-z plus R-r plus R-z               +             +

* In a previous publication (MCCLINTOCK 1938) it was shown that Df-t was not transmitted
through the pollen even when the deficiency was covered by a ring chromosome. Since this
publication, a strain was obtained which will transmit the Df-r chromosome through the pollen
when a covering ring chromosome is present.

chromomeres I to 9 are completely inviable; tissues homozygous deficient
for chromomeres 5 to 9 are likewise inviable. In contrast, the tissues which
are homozygous deficient for chromomeres I to 4 are viable, but the growlh
capacity of these cells is very poor. Only very minute sectors of such hcms-
zygous deficient tissues are found. (See MCCLINTOCK 1938, text fig. 31-38
and this paper, a, fig. 3.) The cells in these sectors are small. The lignified
walls of cells with a full genomic complement are white, but the walls of
cells homozygous deficient for chromomeres I to 4 are brown in color. There
are no well developed plastids in these cells; therefore, the tissues of the
leaf and stalk which are homozygous deficient for chromomeres I to 4 a e
not green and contrast strikingly with non-deficient tissues which ore
green. On exposure to direct sunlight, these cells soon die, and the

FIGURE S.-a. Photograph of a part of a leaf of a Df-r/Df-z plant with ring chromosomes R-t
and R-z. h sector showing the character of the blotch mutant is evident slightly to the right of
the letter a. The wide band to the left of this letter is the mid-rib. The continuous parallel fine lines
are the veins of the leaf. The shorter, narrower lines between these vein& are the sectors of tissue
homozygous deficient for regions I to 4 of chromosome 5 which arise following somatic loss of
both the R-r and the R-2 ring chromosomes. b. Photograph of part of a leaf of a Df-a/Df-2 plant
with a two ring chromosomes, a normal R-z ring and the brown-blotch-dries 2 ring. Two sectors
showing the blotch character are visible to the right and left of the mid-rib, respectively. The
photograph was taken before the tissue had commenced to disintegrate and dry. c. Similar to b.
.I wide sector showing the mutant character blotch-dries is present at each edge of the leaf. The
disintegration and drying process has commenced at the outer edge of each sector. d. Leaves from
plants of the constitution Df-t/Df-2 plus one R-2 ring chromosome. Note the solid (non-varie-
gated) sector of mutant tissue (pink) in each leaf.

FIGURE 4.-Appearance of a young Df-z./Df-a plant with two R-2 ring chromosomes, a normal
R-2 ring and the brown-pink I ring, The plant is variegated for the compound mutant character
brown and pink (white sectors in the photograph). These sectors arise following somatic loss of
the normal R-2 ring.


a

b

C


.:

02 * .           Er' 4        E2

.
k  o                                   ?

   ?


HOMOZYGOUS DEFICIENCIES IN MAIZE
         TABLE 3

549

                                 APPEARANCEOF TISSUES FOLLOWING
CHROMOSOME CONSTITUTION                SOMATIC LOSS OF RING CHROMOSOMES
     OFPLANTS          APPEARANCEOP     INDICATEDINTHE COLUMNS

TISSUES WITH                          -

COMPLETE                   I R-I I R-r

  DEFICIENT        RING
ROD-CHROMOSOME   CHROMOSOMES   COMPLl:MEFT I R-r z R-r r R-z 2 R-z plus  plus
                                                 I R-z z R-s

Df-r/Df-r        I R-I         + -
Df-r/Df-r        2 R-r         +     + -
Df-r/Df-2       I R-2        +                -
Df-r/Df-2   I R-I plus I R-2     +      +       +        -
Df-r/Df-z       2 R-z        +             + -
Df-z,`Df-z       2 R-2        +            + 0
Df-n/Df-a   I R-I plus 2 R-2      +     +       + 0 4 0

DESCRIPTIONOFFIGL-RESFORTHEPLATE

All photographs are of pachytene configurations in microsporocytes. All magnifications are
approximately IIOOX except D 2, E z, and G 2, which are approximately 2750X.
FIGURE A.-Synaptic association of two R-2 ring chromosomes.
FIGURE B.-The lower arrow points to a collapsed normal R-2 ring. The upper arrow points
to a collapsed brown-pink II ring chromosome which is approximately double the size of the nor-
mal R-2 ring.

FIGURE C.-Two collapsed ring chromosomes associated at their centromeres. The arrow
points to the associated centromeres. The normal R-2 ring is below the arrow. Above is the re-
duced ring chromosome which produces the brown Z phenotype.
FIGURE D I.--TWO collapsed ring chromosomes associated at their centromeres. The arrow
points to the associated centromeres. The normal four chromomere R-r ring (above) is slightly
out of focus. The ring below the arrow is a reduced R-2 ring composed of seven chromomeres.
It gives rise to the brown-pink IV phenotype. D 2. Enlargement of the same.
FKGURE; E I .-TWO ring chromosomes associated at their centromeres. The arrow points to the
associated centromeres. The normal, four chromomere R-r ring lies below the arrow. An R-z ring
reduced to three chromomeres lies above the arrow. This latter ring gives rise to the broxn-blotch-
dries ZZ phenotype. E 2. Enlargement of the same.

FIGURES F and H.-The arrows point to the collapsed ring chromosomes in two sister cells of a
Df-r/Df-2 plant nith one R-2 ring chromosome. ;\lI surrounding cells possessed a normal R-2
ring. In one of these sister cells, fig. F, the R-2 ring chromosome tvas enlarged. In the other cell,
fig. H, the R-2 ring chromosome was reduced to two chromomeres. In chromatin constitution the
two rings are equivalent to two normal R-2 ring chromosomes.
FIGURE G I .--A collapsed normal R-2 ring chromosome (lower arro\v) and an R-2 ring chromo-
some reduced to two chromomeres (upper arrow). This latter ring chromosome gives rise to the
compound mutant brown-light green-poor growth. G 2. Enlargement of the same showing this
latter reduced ring chromosome.

FIGURE I.-The arrow points to the collapsed double-sized R-2 ring chromosome which gives
rise to the brown-pink ZZ phenotype.

FIGURE J.-Pachytene configuration in a plant with three ring chromosome. .1 collapsed nor-
mal R-2 ring lies to the left. A normal R-I ring lies immediately above it. The arrow points to the
reduced R-z ring chromosome which produces the brown-light green-poor growth phenotype.


5 50                  II.\KH.iK.\ .\lc(`1,1S1`OC`Ii
tissue disintegrates and dries. Because cells and tissues completely deti-
Gent for these four chromomeres not only are viable but also show pheno-
typic motlitications. it is expected that tissues deficient for segments within
this region woultl not only be viable, but could have much better growth
rates and might show specitic characters.

Studies of somatic mitosis have shown that the larger of the two ring
chromosomes (K-2) produces a double-sized, dicentric ring chromosome
(Prophase. fig. I) approximately once in every hundred nuclear divisions.
X11 of the other nuclear divisions are completely normal; the ring chromo-
some divides normally, the two free sister halves pass to opposite poles
along with the rod chromosomes. In nearly all cases, when a double-sized
ring chromosome is formed at prophase, it remains in the region of the
equatorial plate during anaphase and telophase and is thus completely
eliminated from the sister telophase nuclei. Very occasionally, however, a
double-sized dicentric R-2 ring chromosome becomes broken at anaphase,
and a segment of the broken double-sized ring chromosome enters each
sister telophase nucleus. In both telophase nuclei fusion of broken ends oc-
curs, reestablishing a ring-shaped chromosome, but the content of the ring-
shaped chromosome is usually altered. Similar studies of the smaller ring
chromosome (R-I) have shown that aberrant anaphase behavior occurs
approximately once in every 600 nuclear divisions. In this case, due to its
small size, the ring chromosome is nearly always lost to the sister telo-
phase nuclei. Only rarely have altered R-I rings been detected. The altera-
tions in the chromatin content of the ring chromosomes to be described in
this paper involve the R-2 ring.

DETECTION OF AI.TERED RIKG-SHAPED (`HROMOSOMES 13T MEANS OF
    MC'TANT SECTORS IN THE DF- I , DF- I, DF- I, `DF-2
            AND DF-2 `DF-2 PI.r\NTS

As stated in the previous section. alterations of the R-2 ring are consicl-
erably more frequent than alterations of the R-1 ring. Consequently,
plants with one or two R-2 rings have shown the greatest number of de-
tectable alterations. The plants most suitable for this study have had the
following constitutions: I)f-I II-2 plus one R-I ring: Df-r Df-2 plus two
K-2 rings; IIf- L)f-2 plus two K-2 rings.

In plants of the constitution Df-r Df-2 plus one R-2 ring, a solid sector
of modified tissue should appear following an alteration in the ring chromo-
some which produces a visible change in the character of the tissues. The
size of the sector would depend upon when the altered ring chromosome
arises in the development of the plant and the subsequent rate of growthof
the cells possessing this ring. If the alteration occurred in a single nuclear
division early in the development of the plant, a wide mutant sector could


             NOXluLYC;OUS I)I~1~I~`lI~sc'Il+s IS Jl.iIZl<           5j1
result. If it occurred in a nuclear division late in development, only a small
mutant sector would be pro(lucetl. In l)f-r, Df-2 plants with two ring
chromosomes (either one K-I plus one R-2 or two K-2) or in Df-2,,Df-2
plus two R-2 rings, both solid sectors and sectors which are variegated for
the mutant characters should be produced. The solid sectors should arise
in the tissues of the plant which have only a single ring chromosome-
that is, subsequent to a mitotic loss of one of the ring chromosomes. i\n
alteration in the remaining ring chromosome could give a solid sector with
a modified appearance. 1.ariegated mutant sectors should arise when two
rings are initially present in the tissue, one of which has become altered in
an earlier mitotic division. When the two rings are present, a deficiency in
the altered ring chromosome would bc covered by the presence of this seg-
ment in the normal ring chromosome. Xv0 character caused by the dcfi-
ciency in the altered ring chromosome could appear until after the normal
ring chromosome has become lost in a later nuclear division. .A11 the cells
which arise from a cell which has lost the normal ring chromosome would
be able to show the mutant character associated with the deticiency in the
altered ring chromosome. Since loss of the normal ring chromosome shoultl
occur independently in a number of cells, a sector which is variegated for
the mutant character should be produced. The appearance oi a solid sector
in a plant with a single ring chromosome is illustrated in a and d of figure
3; the appearance of variegated sectors in two-ring plants is illustrated in
b and c of figure 3 and in figure 4.
If a particular mutant character is associated with loss of a particular re-
gion from the ring chromosome- that is, if the character is produced by a
homozygous minute deficiency-- only a restricted number of mutant char-
acters should appear in the Df- 1 Df- I and Df-I, Df-z plants. The segment
within which homozygous deficiencies may be detected is only four chro-
momeres long (fig. 2). If this theory is correct, the same mutant charac-
ters which appear in the Df-I,/Df-1 and Df-I,/Df-a plants should likewise
appear in the Df-2,/Df-2 plants because homozygous deficiencies within
regions I to 4 (chromomeres I to 4, fig. 2) may be produced in all three
types of plants. However, other mutants which are not found in the
Df-I,`Df-r and Df-1, Df-2 plants should be present in the Df-2 `Df-2
plants. These characters should be' related to homozygous deficiencies
within the region -5 to 9 (chromomeres 5 to 9, fig. 2). Such mutant characters
should not appear in the Df-r Df-1 or Df-1, Df-2 plants because the Df-1
chromosome possesses chromomeres 5 to 9* and thus 110 homozygous de-
ficiencies can be produced within this region following aberrations of the
ring chromosome. These anticipations have been completely fulfilled. Since
the altered ring chromosomes may be isolht ed following the formation of
gametes with a deficient rod chromosome and an altered ring chromosome.


5j2                 RARH.~R,\ McCLIN1`O(`K
it has been possible to conduct tests which show that a certain group of char-
acters is associated with region I to 4 and another group of characters
with region 5 to 9.

The frequency of mutant sectors is greater in the Df-rjDf-2 plants
with one or two R-z rings than in the Df-z/Df-z plus two R-2 rings. The
evidence, which is conclusive but which cannot be considered here because
of limitations in space, indicates that this is due to changes in the R-2
ring chromosomes which have deleted rather large segments from the ring.
In a Df-a/Df-a plant, the cells with such large homozygous deficiencies are
either inviable or produce tissues with such poor growth rates that they
are not suitable for precise character studies. However, removal of a large
segment from the R-2 ring in a Df-rjDf-2 plant may result in quite viable
tissues with readily identifiable mutant characters. As an example, re-
moval of region 4 to 8 in the R-2 ring in a Df-a/Df-2 plant would result
in cells homozygous deficient for this region. These cells are inviable. In
a Df-r/Df-2 plant, only region 4 could be homozygous deficient because
the segment j to 8 covering the deficiency in the ring chromosome is pre-
sent in the Df-I rod chromosome.

Considering the number of mitoses taking place in the development of a
plant, the occurrence of an abnormal mitosis which results in an altered
ring chromosome is extremely rare. Once an altered ring chromosome has
been isolated, it can be maintained unchanged through as many plant gen-
erations as desired by avoiding those plants in which this ring chromosome
has again become altered. There are relatively few such plants.

MUTANT CHARACTERS ASSOCIATED WITH REG'ION I TO 4 OF CHROMOSOME j

As stated previously, tissues which are homozygous deficient for the seg-
ments I to 4 of chromosome s-the full extent of the deficiency in the Df-1
rod chromosome-are capable of multiplication but at a very slow rate.
Consequently, only minute visible sectors are produced following com-
plete loss of the ring chromosomes in Df-r/Df-1 or Df-r/Df-a plants. The
mutant characters shown by these homozygous deficient tissues are:
brown cell walls, colorless plastids, very poor growth capacity, and dis-
integration and drying of the tissue when exposed to direct sunlight. It is
to be expected that deficiencies for minute segments within this region
should be viable. They may possess very much better growth capacities
and could show mutant characters. Thus, relatively wide sectors showing
mutant characters associated with a minute homozygous deficiency within
region I to 4 might be produced in these plants. In the Df-r,`Df-r or
Df-r/Df-2 plants, all the mutant characters observed as sectorials
should be related to changes within region I to 4 of chromosome 5. In the
Df-z/Df-a plants, these same types of mutant characters should appear


           HOMOZYGOUS DEFICIENCIES IN MAIZE          553
plus additional mutant characters which are not found in the former types
of plants. These latter mutants should be related to changes within region
5 to 9-
The various types of mutant characters having a clearly recognizable
phenotypic expression which have appeared as sectorials in all three types
of plants-that is, those mutants associated with region I to 4 of chromo-
some j-have been divided into three groups: (I) Those which show a
single recognizable character in the mutant sector (simple mutants); the
tissue may have a normal growth rate. (2) Those which exhibit two or
more of these characters in a single sector (compound mutants); the tis-
sues may have a normal growth rate. (3) Sectors with distinct mutant char-
acters. These sectors are always narrow-that is, associated with a reduced
growth rate of the cells of the sector; many of these are recognizable as
compound mutants.
The simple mutants to be described involve some obvious color change.
Although other mutants would be expected and are probably present,
their positive detection as sectorials in these plants is both difficult and
uncertain. Therefore no attempt was made to isolate mutants which did
not produce an unmistakable character change which could express itself
as a sector in a plant. The color mutants belong to the unmistakable class.
  Group I. Simple mutants which may haee a ,xormal gro-wth rate
                Brown cell walls
The lignified cell walls are brown in color. This character is similar in all
details to the character produced by the recessive mutant brown mid-rib
(symbol, bm) previously located within the region I to 4 of a normal rod
chromosome 5. Henceforth the character will be designated as brown. The
normal R-I and R-2 rings are known to carry the dominant allele of bm
(Bm colorless cell walls). (For a complete description of the bm phenotype
and proof of the presence of bm in the normal ring chromosomes, see Mc-
CLINTOCK rg$3.) Three different ring chromosomes producing the brown
phenotype have been isolated. Two appear to be normal in size and could
not be distinguished from normal R-2 rings. Either ring will produce the
brown character in Df-r/Df-I, Df-r/Df-a or Df-a/Df-2 plants. The third
brown ring is reduced to six chromomeres (upper collapsed ring, fig. c,
Plate I). When this ring is present, brown will appear only in Df-r/Df-r or
Df-r/Df-a plants. In Df-a/Df-a plants the cells which have only this ring
are inviable. In this case, the three chromomere deficiency in the ring chro-
mosome probably includes a segment of region 5 to 9 which could express
itself as a homozygous deficiency only in the Df-s/Df-2 plants. As ex-
plained previously, in the Df-r,`Df-r or Df-r/Df-a plants, the cells with
only this reduced ring chromosome might possess only a minute homo-


.i 54                  I1.\KIS.\K.\ 1l~~~`I.IN'I`OC'K
zygous deticiency and thus be quite viable. This would explain the differ-
ence in viability of the cells possessing only this ring in the three types of
plants.                 Pink
Chlorophyll is absent although plastids are present. In the stalk, the
number of layers of cells from the outer circle of bundles to the epidermis
frequently is greater in the pink sectors than in the non-modified parts of
the plant. Consequently, the pink sector produces a protuberance. On the
stalk and in the upper leaves of the growing plant, the color is an intense
salmon pink. ;\s the leaves mature. the pink color gradually fades to white.
Two ring chromosomes giving rise to the pink phenotype have been iso-
lated, but the constitution of only one of these rings has been investigated
cytologically. It is an R-2 ring with no clearly observable change in chro-
matin constitution. It produces the pink phenotype in either a Df-I/Df-I,
a Df-I, Df-2 or a Df-2 llf-2 plant.

                    Blotch
The chlorophyll pattern is finely speckled (a, fig. 3). So ring chromosome
producing a blotch character associated with a good growth capacity
has been isolated so far, although a large number of such sectorials have
been observed in both Df-r/`Df-a and Df-a/Df-2 plants. Several ring chro-
mosomes producing blotch but associated with poor growth capacities
have been isolated. Sone of these has been investigated extensively.

                  Blotch-dries
This character is similar to blotch although the chlorophyll color usually
is less intense. In the young leaf, the character is first detected as a light
green sector. The blotch pattern of the chlorophyll is acquired as the leaf
matures. After a short time, the cells in the sector which have been exposed
to direct sunlight begin to disintegrate. Finally, the whole sector so exposed
becomes a mass of dead. dried tissue (see b and c. fig. 3). Two ring chromo-
somes producing blotch-dries have been isolated recently. Seither has
been sutticiently investigated as yet.
  Gro1rp -3. C'onzponnd wtctartts zifhich mily hw wormd groih rates
              Brown-pink
Many sectors showing the combined characters of brown and pink have
been observed in all three types of plants. Four different ring chromosomes
producing this compound character have been isolated. One is an obvi-
ously reduced R-2 ring chromosome composed of seven chromomeres
(lower collapsed ring, fig. 1). Plate I). Two are slightly reduced R-2 rings.
and one is an enlarged K-Z ring. approximately double the size of the nor-
mal R-Z ring icollapsetl ring. upper arrow. tig. H and fig I. Plate I). The


             HOJIWYGOUS I)I~FIClESCII:S IN M.\I%I.:             ---
                                                           333
brown-pink character produced by the seven-chromomere ring appears
only in the Df-I/Df-1 or Df-1 Df-z plants. In the Df-2:Df-2 plants, the
cells with only this ring chromosome are inviable. The other three ring
chromosomes will produce the brown-pink character when present in either
a Df-r,`Df-1. a Df-I, Df-2, or in a Df-2, Df-2 plant.
            Brown-pink-dries
.\t the initial stage, the sectors showing this compound mutant charac-
ter arc strictly comparable to the compound mutant brown-pink. However.
as in blotch-dries, on exposure to sunlight, the cells die and the tissue dries.
These sectors appear in both Df-r Df-2 and Df-2, Df-2 plants. Several
rings causing this compound mutant character have been isolated, but
none has been investigated fully.

               Brown-blotch-dries
The sectors exhibiting this compound mutant are identical in appearance
and behavior to the blotch-dries sectors described above. In addition, horv-
ever, the lignitied cell walls are brown. Two ring chromosomes producing
this compound mutant have been isolated. One is a slightly enlarged R-2
ring. The other is a much reduced ring chromosome composed of only
three chromomeres (ring above arrow, fig. E, Plate I). When the former
ring chromosome is present, the character will appear in either a Df-1 Df-2
or a Df-2, Df-2 plant. When the latter ring is present, the compound
mutant character will appear only in a IIf- :D~-I or a Df-I, Df-a plant.
In the Df-2i/Df-2 plants, the cells which possess only this ring chromosome
are inviable-probably because of the six chromomere deliciency.
      &OUP 3. C'o~?~po~t~~d ntuttin ts ,Z~IIOS~ groWth rtitfs
           tzre considerably reduced
         Brown-pink-dries-poor growth
The sectors showing this group of characters are similar in appearance to
the compound mutant brown-pink-dries of group 2. However, the growth
rate of the mutant'tissues is always considerably reduced. Thus, only nar-
row sectors are formed. Three rings producing this compound mutation
have been isolated, but only one has been sufficiently studied. It is a
slightly reduced R-z ring. The character which this ring produces is ex-
actly the same in the Df-I/Df-1, Df-r/Df-2, and Df-a/Df-2 plants. Thus,
the phenotypic modification which this ring chromosome induces is prob-
ably restricted to a segment within region I to 4 of chromosome 5.
          Light green-poor growth
The chlorophyll color in these sectors is a light green. but the growth
rate of the cells is so reduced that sectors are not wider than ,: or 4 mm.


556                  BARBAR. McCLINTOCK
Only one ring chromosome producing this character has been isolated. It
possesses only two chromomeres. Thus, the character associated with this
ring appears only in the Df-I,`Df-1 and Df-I,`Df-2 plants. It does not ap-
pear in the Df-z/Df-2 plants due to the extensive deficiency producing in-
viability in the cells possessing only this ring.

Brown-light green-poor growth

Sectors showing this character are similar in appearance to those de-
scribed above. In addition, the lignified cell walls are brown. Only one ring
chromosome giving rise to this compound mutant has been isolated. It
possesses two chromomeres (upper arrow, fig. G I and enlargement, fig.
G 2, Plate I). Due to the extensive deficiency in this ring chromosome, the
characters associated with this ring appear only in Df-I/Df-1 or Df-I/Df-2
plants.

Many sectors exhibiting very poor growth capacities associated with a
blotched chlorophyll pattern or with colorless tissues, either with or with-
out brown,have been observed in the Df-I/Df-I,Df-I:`Df-2, and Df-a/Df-2
plants. No attempt has been made to isolate the rings responsible for such
characters because of the impracticability of work with characters having
such poor growth rates.

MUTANT CHARACTERS ASSOCIATED Vv'ITH REGION

j TO 9 OF CHROMOSOME j

The mutant characters which may be associated with alterations in the
R-z chromosome within region j to 9 are those which do not appear in the
Df-I/Df-1 or Df-I,jDf-2 plants but appear only in the Df-a/Df-2 plants.
It is difficult to obtain sufficient seed for an extensive investigation of the
types of mutants appearing in this latter type of plant. Only one to several
kernels are likely to occur on an ear of a Df-n/Df-2 plant, since the Df-2
chromosome is not transmitted through the female gametophyte without
the compensating R-2 ring chromosome. During meiosis, the R-2 rings are
so frequently eliminated that few megaspores are formed possessing this
ring, and thus few seed may develop. Among the several thousands of
plants which have been examined, a number of mutant sectorials have been
observed. Many of these are related to changes in region I to 4 of the R-2
ring chromosome. Only a few clear-cut mutants have been observed which
may be related to changes within region 5 to 9. In these plants, many mu-
tant sectorials are composed of tissues which grow too poorly to be useful
for character studies. Ko attempt has been made to isolate the changed
rings responsible for the mutations with very poor growth rates. When a
whole plant or half of a plant was variegated for such a character, a cyto-
logical study was made to determine the types of ring chromosomes which


           HOMOZYGOUS DEWICIENCIES IN MAIZE          557
were present. In all examined cases, one of the ring chromosomes was de-
cidedly reduced. Following loss of the normal ring chromosome to some
cells during development, a relatively large homozygous deficiency had to
be present. If these cells reproduce at a very reduced rate, only very
narrow sectors of homozygous deficient tissue could be formed. The pres-
ence of a reduced ring in all examined plants showing variegated sectors
with considerably reduced growth rates leads one to suspect that the cells
of these sectors possess only the reduced ring chromosomes-that is, they
are homozygous deficient for the segment which had been deleted from the
ring chromosome.
Of the mutants which possess a good growth capacity, only three have
been found which may be ascribed to region 5 to 9; and up to the present
time, only two of these have been proven to be in this region. These mu-
tants are:
              Pale green
The chlorophyll color in the leaves and the stalk is a pale green. The pale
green character does not develop in the very young plants but becomes
very striking in the older plants. The growth rate of this tissue is usually
quite normal. Only one pale green ring has been isolated. No obvious
change in the chromatin constitution of this R-2 ring was observed.

                   Striate
The sectors showing this character are composed of streaks of yellow-
green tissues. Only one ring chromosome has been isolated which produces
this character. This R-2 ring is slightly reduced. A gamete with the Df-2
rod chromosome and the R-2 striate ring chromosome is transmitted
through the female gametophyte but not through the pollen. The location
of the character within region 5 to 9 has been determined by use of a third
deficiency rod chromosome (Df-3) possessing a small deficiency located
within the limits of region 5 to 9. Striate appears when the striate ring
chromosome is present in Df-P,/Df-g plants.

                   White
Sectors with a chalk white color have been observed in the Df-a/Df-2
plants but ring chromosomes producing white have not been isolated as
yet. Since similar sectors have not been observed in many thousands of
Df-r/Df-2 plants, it is assumed, although not proven, that this character
is related to an alteration within region 5 to 9.
The pale green ring is the only one of this series which has been exten-
sively investigated. It is not related to striate, since the pale green charac-
ter does not appear in Df-a/Df-3 plants, whereas striate does appear in
these plants. The following tests have shown that the pale green character


558                  B.iHBhK.1 McCI,INTOCK
is related to region j to 9 and not to region I to 4 of chromosome 5. Follow-
ing isolation of the pale green ring, plants were obtained with the constitu-
tion Df-r/Df-a plus the pale green ring. These plants were indistinguish-
able from those of a similar constitution but having a normal R-2 ring.Thus,
pale green does not appear in a Df-r background. The functional male
gametes produced by these plants possess the Df-z rod chromosome plus
the pale green ring chromosome. When pollen of such plants was placed on
silks of plants of the constitution Df-z/Df-a plus two normal R-2 ring chro-
mosomes, all the resulting individuals (Df-a/Df-a plus one normal R-2
ring plus the pale green R-2 ring) were variegated for pale green. The varie-
gation arose as a consequence of the mitotic losses of the normal R-2 ring
chromosome contributed by the female parent, thus giving rise to tissues
containing only the pale green ring chromosome. It is in these tissues that
the pale green character appears.

CYTOLOGICAL OBSERVATIONS OF THE RING-SHAPED CHROMOSOMES
     WHICH GIVE RISE TO THE MUTANT CHARACTERS

A pachytene configuration showing the association of two normal R-2
ring chromosomes is shown in figure A, Plate I. When a single R-2 ring
chromosome is present or when two R-2 ring chromosomes which have
failed to associate at the meiotic prophase are present, the ring chromo-
some is collapsed (lower arrow, fig. B and G, Plate I ; ring below arrow, fig.
C, Plate I). This results from non-homologous associations of the chro-
matin forming the ring (MCCL~NTOCK 1933~ 1938).

Following the formation of a double-sized dicentric ring chromosome in
a mitotic prophase and its breakage in the following anaphase, an enlarged
ring chromosome may be formed in one telophase nucleus and a reduced
ring chromosome in the sister telophase nucleus (see fig. I). This situation
is illustrated by the constitution of the ring chromosome in two sister cells
(arrows fig. F and H, Plate I). These two sister cells are in the prophase
of meiosis; therefore the event which resulted in the formation of these
two aItered ring chromosomes occurred at the last premeiotic anaphase.
In individual cases, R-2 rings have been observed with various degrees of
reduction in chromatin constitution. These range from a barely observable
deficiency in the ring chromosome to deficiencies so large that the ring
chromosome was composed of but a single large chromomere. Likewise, in
individual cases, R-2 ring chromosomes have been observed with dupli-
tated segments varying in extent from a very small duplication to one
which produced a ring chromosome eight times the size of the normal
R-2 ring.

Cytological observations of the composition of the ring chromosomes
which produce visible mutations are summarized in table 4. As stated pre-


             HOMO%\-GOUS DEFICIENCIES IN M.l\IZE          559
viously in the description of the mutant characters, several ring chromo-
somes have been isolated which produce the same mutant character (simple
mutant) or the same group of mutant characters (compound mutant).
Although the character which each of these ring chromosomes produces
appears to be the same, the event which is responsible for the character
which the ring chromosome produces occurred independently in each case
in a single Df-r,`Df-1, Df-r/Df-2, or Df-z/Df-a plant. The numerals I, ZZ,
and III, etc., have been used to distinguish ring chromosomes producing
the same character or the same group of characters which have originated
independently of one another. For example, brown I was isolated from a
sector of a Df-r/;Df-2 plant, brown ZZ from a sector of another Df-r/Df-a
plant, and brown ZZZ from a Df-2,`Df-2 plant. Where a photographic illus-
tration of a particul!$r altered ring chromosome is reproduced in this paper,
the figure reference is indicated in the table. Four of the fifteen ring
chromosomes showed no observable change in the constitution of the R-2
ring. All four ring chromosomes are associated with simple mutants (brown
ZZ, brown ZZZ, pink, pale green). In nine cases, the ring chromosome was
reduced. In two cases, the ring chromosome was enlarged (brown-pink ZZ,
brown-blotch-dries I). The brown-pink ZZ ring, when first observed, was
approximately double the size of the normal R-2 ring (upper arrow fig. B
and fig. I, Plate I). The brown-blotch-dries Z ring is only slightly enlarged.
In both of these cases, the alteration in the ring chromosome, which is re-
sponsible for the character the ring chromosome produces, could have oc-
curred subsequent to a previous alteration which had enlarged the R-2
ring chromosome. It is likewise conceivable that the enlargement of the
ring chromosome could have occurred following the alteration which pro-
duces the character but before isolation of the ring. It is known that the
enlargement itself is not the factor responsible for the appearance of the
character which such an enlarged ring chromosome produces. This is clearly
indicated by the enlarged brown-pink II ring. This ring, when first isolated,
was approximately double the size of a normal R-2 ring chromosome. Be-
cause of the increased length of the chromonema of this ring chromosome,
aberrant mitoses involving this ring chromosome occur with increased fre-
quency. Furthermore, the added length of the chromonema of this ring
chromosome brings it into the range where aberrant mitoses may fre-
quently lead to recoverable alterations in the ring chromosome. Because
of its size, it was expected that this ring chromosome would not maintain
itself but would tend to become reduced to the size where aberrant mitoses
would lead most frequently to elimination of the ring chromosome rather
than alterations in its chromatin content. This proved to be true. Several
strains have been isolated in which this ring chromosome is only slightly
larger than the normal R-z ring chromosome. At this reduced size! the


560                 B;\RBARA &fcCLINTOCK
ring chromosome tends to remain stable because aberrant mitoses now
tend to eliminate the ring rather than alter its chromatin composition. It
should be emphasized, however, that the characters produced by the
brown-pink II ring following its reduction in size from a double-sized ring
to one only slightly larger than a normal R-2 ring have not been altered by
the reduction. It is assumed that even in its enlarged state, the brown-pink
II ring chromosome possessed a minute deficiency which was responsible
for the compound mutant character it produced. No duplications of the
remaining segments of the ring chromosome or subsequent deletions of
these duplicated segments should alter the expression of the character, for
the minute deficiency would still be present regardless of the presence or
absence of these duplicated segments.

Chromalin consliluliox of Ihe ring chromosomes which produce mnl~tartt characlers.

I. So observed change in constitution of the R-z ring.
  brown II; brown III; pink; pale green.
2. Slightly reduced R-2 ring: Loss of approximately one chromomere.
  brown-pink I; brown-pink III; striate.
3. Obviously reduced R-2 ring: Loss of two or three chromomeres.
  brown-pink IV (fig. D, Plate I); brown-pink-dries-poor growth; brown I (fig. C, Plate I).
4. Very reduced ring: Loss of six or seven chromomeres.
  brown-blotch-dries II (fig. E, Plate I); light green-poor growth; brown-light green-poor
  growth (fig. G, Plate I).
5. Enlarged ring: Larger than a normal R-2 ring.
  brown-pink II, approximately double the size of a normal R-2 ring (fig. B and I, Plate I);
  brown-blotch-dries I, approximately + larger than the normal R-z ring.

In II of the 15 cases it has been established that the ring chromosome
has undergone an alteration. It would require considerable space to indi-
cate the methods used which allow one to be certain that the ring chromo-
some being observed in a particular plant is the ring chromosome respon-
sible for the character ascribed to it. The methods will be considered in a
separate report.

TRANSMISSIONS OF ALTERED RING CHROMOSOMES

THROUGH THE MALE AND FEMALE GAMETES

Tests have been made of the transmissions of??ie various altered ring
chromosomes through the female gametophyte with the Df-1 or Df-2 rod
chromosomes and through the pollen with the Df-2 rod chromosome. These
results are summarized in table 5. Transmissions through the pollen of the
various altered ring chromosomes with the Df-1 rod chromosome have not
been included in the table, since a male-transmissible strain of Df-1 has
only recently been obtained, and the tests have not been completed. The
cytological determination of the type of ring chromosome associated with


           HOMOZYGOUS DEFICIENCIES IN Mr\IZE        561
each particular mutant has been included in column 2 of this table, since
it allows the tests to be more readily interpreted. Since many of these
altered ring chromosomes are readily transmitted through the pollen, even
in competition with gametes carrying normal ring chromosomes, it has
been po@ble to obtain various combinations of the ring chromosomes in
Df-r/`Df-1, Df-r/Df-a, and Df-2/Df-2 plants. These will be described in
the following section.

                   TABLE 5
Cametic Iransmissiows oj the altered ring chromosomes with the Of-1 or Df-2 rod chromosomes.
Transmissions of the altered ring chromosomes with Ihe Df-r rod chromosome lhrotrgh the pollen have
not been inclrlded in the table because these tests have not been completed. + represents transmission.
o repmenls no lransmission. - indicates the lcsl has not heen completed.

TR.4SSMISSIOKS TRANSYISSIOX

    RIXG CHROMOSOME       CHROMATIS CONSTITVTIOS OF   THROUGH 0   THROUGH a"
                      ALTERED R-z RISC         WITH :       WITH
                                       Df-r   Df-2     I>f-2
                                                            -
brown I              Reduced to six chromomeres    +
brovi-n II            No apparent reduction   + 1   1
hro\vn III           pie apparent reduction       + +      +
pink             No apparent reduction       + +     +
pale green           No apparent reduction      + +     +
striate            Slightly reduced            -    +
brown-pink I         Slightly reduced          + +   i
brown-pink II         Enlarged             + +      +
brown-pink III        Slightly reduced          + +      +
brown-pink IV          Reduced to seven chromomeres   +
brown-pink-dries-poor growth  Slightly reduced     + i       Cl

brown-blotch-dries I      Slightly enlarged          + +   J
brown-blotch-dries II       Reduced to three chromomeres  +     0       0
light green-poor growth      Reduced to two chromomeres   +     0       0
brown-light green-poor growth Reduced to two chromomeres   +     0       0

THE CHARACTERS PRODUCED IN PLANTS FOLLOWING COMBINATIONS

OF ALTERED RING CHROMOSOMES: PROOF THAT THE COMPOUND
      MUTANTS ARE COMPOSED OF TWO OR MORE

OF THE SIMPLE MUTANTS

It is obvious from the description given earlier in this paper that the
characters exhibited bythe compound mutants bear a relation to the simple
mutants. The compound mutants appear to be composed of a combination
of two or more of the observed simple mutants. It was also obvious in
this study that the sectorials showing compound mutant characters oc-
curred far more frequently than those showing a simple mutant character.
Sectorials showing a simple mutant character were relatively rare. The
method by which ring chromosomes become reduced in size at a mitotic
anaphase would suggest that adjacent blocks of chromatin should fre-


sh2                 B,lRB.iRA McCLINTOCK
quently be deleted from the ring chromosome following an aberrant mitosis
which results in an alteration in the ring chromosome. If each of the simple
mutants is produced following loss of a particular minute segment from the
ring chromosome, then the compound mutants should be produced follow-
ing loss of two or more of these segments. It would be expected that break-
age of the double-sized ring at a mitotic anaphase would delete a block of
chromatin more frequently than a very minute segment. Thus, compound
mutants should appear far more frequently than simple mutants. With
this interpretation, one should be able to deduce the order of the various
segments in the ring chromosome which produce a particular character
when homozygous deficient, merely by observing the characters which are
present in the compound mutants. For illustrdtive purposes, one may use
the characters pink, brown, and blotch dries. The most frequent types of
compound mutations involving these characters are: brown-pink, brown-
pink-dries, and brown-blotch-dries. Since brown may be with either pink
or blotch dries, the suggested order is (I) pink followed by (2) brown fol-
lowed by (3) blotch dries. Loss of the first two adjacent segments would
give rise to the compound mutant brown-pink; loss of the second and third
adjacent segments would give rise to the compound mutant brown-blotch
dries; loss of all three adjacent segments would give rise to the compound
mutant brown-pink-dries.
Proof that the compound mutants are composed of two or more of the
simple mutants may be obtained readily. It is known that all of the mutant
types behave as recessives. It is only necessary to combine two altered rings
each of which gives a character or a group of characters in a Df-r/Df-2 or a
Df-z/Df-2 plant and observe the characters of the tissues which possess
the two altered ring chromosomes. The method is essentially that which is
diagrammed in table I, and the results are in complete agreement with it.
Various combinations of the altered ring chromosomes have been made.
Those which involve the mutants brown, pink, brown-pink, brown-pink-
dries, brown-blotch-dries are given in table 6. The appearance of the cells
and tissues possessing both ring chromosomes are given in column 2. In
column 3, the character of the tissues which possess only one or the other
of the altered ring chromosomes (following mitotic loss of one or the other
ring chromosome, respectively) has been indicated. Where more than one
similar combination was made, the particular altered ring chromosome
(that is, I, II, or III) is given below the character in each specific combina-
tion. It made no difference which ring chromosome producing a given mu-
tant type was used. In every tested case the results were the same. Com-
bination I of table 6 (X plus Xormal) gives the characters resulting from
the combination of any one altered ring chromosome, X, with a n&ma1 R-2
ring chromosome. In all of the 15 isolated cases the normal R-2 ring chro-


             HOMOZYGOUS DEFICIENCIES IN M;\IZE        563
mosome suppressed the character produced by the altered ring chromosome
when both were present in the same cell. However, the presence of the
altered chromosome in these plants was made obvious by the variegation
which appeared (fig. 4). This variegation resulted from the mitotic losses
of the normal R-2 ring chromosome. The character produced by the altered
ring chromosome could then be expressed.

                          TABLE 6
Phenotypic characters in Df-r/Of-r, 0j-1/Dj- 2, and Df-z/of-z plants with various ccr~~hic~u~ions of
ring chromosomes logelher with the type of uariegalion which is obseroed iu enrh oJ these plants folloti i,lg
somatic loss oj one or the other ring chromosome, respectia~ely. + indirafes norrnul, nor~-mutdd tissue.

I. X' plus normal R-z

+     Loss of normal R-z ring: X tissue.
    Loss of X ring: + tissue.

2. brown II plus brown 111

3. brown II plus pink

Plant totally  Lcxs of brown II ring: So change in tirsue character.
brown    Loss of brown III ring: So change in tissue ch;rractrr.

  +     Loss of brown II ring: pink tissue.
       Loss of pink ring: brown tiswe.

4. brown plus brown-pink
  II =     I
  III "     I
  II y    II
  III =    II

Plant totally  Loss of brown ring: bmwn-pink tissue.
brown    Loss of brawn-pink ring: No change in ti.>ue ch:ir:lcter

5. brown plus brown-blotch-dries
  II *       I
  III y       I
  III y      II

Plant totally  Loss of brown ring: brown-blutch-dries tissue.
brown    Loss of brown-blotch-dries ring: No change in tissue character.

6. brown I plus brown-pink-dries II

Plant totally  Loss of brown ring: brown-pink-dries tissue.
brown    Loss of brown-pink-dries ring: So change in tissue character.

7. pink plus brown-pink I          pinkt

Loss of pink ring: brown-pink tissue.
Loss of brown-pink ring: pie change in tissue character.

8. pink plus brown-blotch-dries I

+    LOSS of pink ring: brown blotch-dries tissue.
    Loss of brown-blotch-dries ring: pink tissue.

Q. brown-pink plus brown-blotch-dries  Plant totally  Loss of brown-pink ring: brown-blotch-dries tissue.
    I `I       I        brown   LOSS of brown-blotch-dries ring. broxtn-pink tiswe.
    I y      II                                          __-

  o X represents any altered ring chromosome producing a mutant character.
  t Due to lack of chlorophyll, plants that are totally pink do not survive beyond the seedling stage. The test was
made by combining these two ring chromosomes with a normal ring chromosome. The character produced by the comb-
nation (column 2) and the characters indicated in column 3 were obtained from sectors of these plants which had Ic)st
the normal ring chromosome.

Combination 2 in table 6 indicates that the simple mutant brown pro-
duced by the brown II ring chromosome is identical with the simple mu-
tant brown produced by the brown III ring chromosome. Plants with
these two ring chromosomes are totally brown. So detectiblc alteration


564                 BARBAR McCLISTOCK
occurs in the tissues following loss of either ring chromosome. Combination
3 indicates that the simple mutant brown II and the simple mutant pink
are completely independent of one another, for the pink character is sup-
pressed in the presence of the brown II ring chromosome, whereas the
brown II character is suppressed in the presence of the pink producing
ring chromosome. However, the plant is variegated for both brown and
pink following mitotic losses of the pink ring or the brown ring, respec-
tively. There were no mutant sectors with the combined character brown-
pink. Combination 4 indicates that the brown in the compound mutants
brown-pink I and brown-pink II are identical with the brown character
produced by either the brown II or the brown III ring chromosome, for
these plants are totally brown although variegated for pink, following
mitotic losses of the brown II or the brown III ring chromosome, respec-
tively. Since the character brown II is identical with brown III, the brown
character in the compound mutant brown-pink I must be identical with
the brown character in the compound mutant brown-pink II. Similarly,
as combination 5 indicates, the brown character of the compound mutants
brown-blotch-dries I and brown-blotch-dries II is identical with the simple
mutant character produced by either the brown II or brown III ring chro-
mosomes. The plants are totally brown but variegated for blotch-dries
following mitotic losses of the brown II or brown III ring chromosomes,
respectively. Combination 6 indicates that the simple mutant brown pro-
duced by the reduced brown I ring chromosome is identical with the
brown produced by the ring chromosome giving the compound mutant
brown-pink-dries II. (This latter compound mutant has not been men-
tioned previously in this paper.) The homology of the simple mutant pink
with the pink produced by the ring chromosome giving the compound
mutant brown-pink I is shown by combination 7. However, the pink ring
covers the brown part of the compound mutant produced by the altered
ring chromosome giving brown-pink I. Combination 8 shows that the
simple mutant pink and the compound mutant brown-blotch-dries I have
no overlapping effects. The pink ring suppresses the brown-blotch-dries I
character. Likewise, the brown-blotch-dries I ring chromosome suppresses
the pink character produced by the pink ring. The combination of the
compound mutants brown-pink I with either brown-blotch-dries I or II
indicates that the brown character is identical in both compound mutants,
for the plants are totally brown. However, the pink character produced
by the brown-pink I ring is suppressed by the brown-blotch-dries rings,
and the blotch-dries character produced by the latter ring chromosomes is
suppressed by the brown-pink I ring chromosome.         f
From the combinations given in table 6, it may be seen that the brown
mutant produced by an altered ring chromosome is the same whether it


             HOMOZYGOUS DEFICIENCIES IN MAIZE        565
appears as a simple mutant or in combination with other mutants. A simi-
lar conclusion may be drawn regarding the pink mutant, although there
is less experimental evidence for this on the basis of the combinations
given in table 6. It is quite obvious from these combinations that brown,
pink, and blotch-dries are independent mutants and that the compound
mutants result from combinations of these simple mutants. As stated ear-
lier one may deduce the order of the segments in the R-2 ring chromosome
which are responsible for the mutant effects. This order, as stated pre-
viously, was: pink followed by brown followed by blotch-dries. The experi-
mental evidence given in table 6 supports this interpretation.
From these tests it may be concluded that the compound mutants are
produced by combinations of two (or more) of the observed simple mu-
tants and that the mutants giving the same phenotypic appearances, al-
though arising independently of one another, are identical.
To discuss adequately the characters which are produced in plants
carrying the deficient rod chromosomes following combinations of the
various altered ring chromosomes would require considerable space and
cannot be undertaken within the limits of this paper. However, the results
of combining an altered ring chromosome with normal, non-deficient rod
chromosomes 5 should be mentioned. If a plant possesses a normal rod
chromosome 5 carrying Bm and likewise any one of the various altered
ring chromosomes, the characters which these ring chromosomes would
produce if deficient chromosomes 5 were present now do not appear. The
plants are normal. However, when the ring chromosome is returned to
plants carrying the deficient rod chromosomes in a successive plant gen-
eration, the character produced by the ring chromosome again appears.
The characters produced by the ring chromosomes are thus completely
recessive to `dominant alleles" which are present in segment I to 9 of the
normal, non-deficient rod chromosome 5. This is not true, however, if a
brown producing ring is combined with normal rod chromosomes 5 carry-
ing the recessive mutant bm. All the ring chromosomes which give rise
to the brown character (simple or compound mutants) in plants with the
deficient rod chromosomes have been placed in plants possessing normal
rod chromosomes 5 carrying bm. In every case, the plants are totally
brown. In these plants, only the brown character appears. The pink,
pink-dries, blotch-dries, or light green characters are suppressed by this
rod chromosome 5. From these combinations it is strikingly evident
that the brown character produced by the altered ring chromosomes is
homologous to the known mutant bm previously located within region I
to 4 of a normal chromosome 5 (MCCLINTOCK 1938). A normal chromo-
some 5 carrying any one of the other mutants produced by the altered
ring chromosomes has not been isolated as yet.


  RE\`Il<\V OF THE EVIDENCE \VHICH INDIC.kTES THAT THE CHXRACTl?KS

    PRODI-CED BY THE ALTERED RING CHROMOSOMES ARE CAT`SED

                BY HOMOZYGOUS DEFICIENCIES
Throughout this paper it has been assumed that the mutant characters
are caused by homozygous minute deficiencies. It has been determined by
many observations that ring chromosomes may increase in size by dupli-
cations and repeat duplications of segments composing the ring, or the)
may become reduced in size through loss of segments from the ring. It has
also been determined that whole plants or sections of a plant which possess
a ring chromosome with duplicated segments are not obviously modified
in phenotypic appearance. Thus, the ring chromosomes with deficiencies
are the ones which would be expected to produce an alteration in the
phenotypic appearance of the tissues in the plants with the deficient rod
chromosomes. Deficient ring chromosomes must be produced in some cells
of some of these plants. It could be objected that such homozygous de-
ficiencies might be expected to be cell lethal, and therefore the character
produced by the altered ring chromosome could be caused by some other
process than deficiency. However, this has proved not to be true. It is
known that cells and tissues homozygous for the four chromomere de&
ciency of the Df-1 rod chromosome are viable. It is also known that these
cells and tissues are decidedly modified. The lignified cell walls are brown.
This brown is strictly comparable in time of development in the cell wall,
in color, and in behavior to light to that produced by the known mutant
bm which had previously been located within the limits of region I to 4
of a normal chromosome 5. (Th e c h aracter of the lignified walls produced
when the altered ring chromosomes brown I, II, and 111, brown-pink 1,
II, etc., are present is likewise strictly comparable to the character pro-
duced by the mutant bm.) Iz-o chlorophyll develops in the cells homozy-
gous for the four chromomere deficiency. Therefore, with regard to chloro-
phyll, these tissues are colorless. Furthermore, on exposure to sunlight,
these cells die and the tissue dries. These tissues have a very reduced
growth capacity. It is known that the characters brow-n, pink, and blotch-
dries must be related to changes within the region I to 4 of chromosome 5
-that is, within the limits of the deficiency in the Df-r chromosome. It
has also been shown that the two very reduced rings (light green-poor
growth and brown-light green-poor growth) produce tissues with a de-
cidedly reduced growth capacity. Since the latter two ring chromosomes
obviously do not have enough chromatin to cover the deficiency in the
Df-1 chromosome, the reduced growth could be ascribed to the presence
of the two chromomere deficiency in each case. If we add together the
characters produced by the changed rings- that is, brown plus pink (no


           HOJIOZ\-GOCS DEFICIENCIES IX Jl,\IZE         567
chlorophyll) plus blotch-dries plus poor growth- we arrive at the com-
pound character brown-colorless-dries-poor growth, which is exactly the
compound character exhibited by the tissues which are homozygous de-
ticient for the four chromomeres deleted from the Df-r chromosome. This
four chromomere deficiency expresses itself as a compound mutant just as
brown-pink, brown-pink-dries, and brown-blotch-dries express themselves
as compound mutants. Evidence for the compound nature of these mu-
tants has been given in this paper (see table 6). It would bc difficult to
arrive at a more simple explanation of all these facts than that the char-
acters are produced as the result of homozygous deficiencies, each indi-
vidual character being produced by a relatively minute deficiency of a
specific locus. This will explain both the simple and the compound mu-
tants: the simple mutants resulting from loss of one specific locus, the
compound mutants resulting from loss of two or more specific loci. It will
also account for the recessive expression of these characters, for it is known
that plants with one normal chromosome 5 and either the Df-r or the Df-a
chromosome 5- that is, hemizygous for regions I to 4 or I to 9, respec-
tively-are phenotypically normal.
Notwithstanding the cytological observations of reduction in size of
many of the ring chromosomes giving mutation effects, genie mutation
might be suggested as an alternative to homozygous deficiencies as the
cause of the mutant characters. If so, two or more genie mutations would
have to occur simultaneously in closely related loci in many instances to
account for the considerably greater frequency of occurrence of the com-
pound mutations. Furthermore, the rate of such mutations in the ring
chromosome would have to be considerably greater than the rate of muta-
tion in the same region of a normal rod-shaped chromosome 5. Our knowl-
edge of "genie" mutations would not lead us to anticipate such behavior,
for we know of no cases where supposedly genie mutations behave in
exactly this manner.
Likewise, one might argue that the alterations in the ring chromosomes
producing mutant effects are due to changes in the relative positions of
the genes in the chromatin of the ring. One must then explain what should
be expected of the reduced ring chromosomes, for they are being produced
in these plants following aberrant mitoses which alter the chromatin con-
tent of the ring. Again, on this basis, one would have to explain away the
cytologically obvious deficiencies in nine of the 13 isolated ring chromo-
somes which produce mutant effects. Since tissues homozygous deficient
for chromomeres I to 4 of chromosome 5 are known to produce a compound
mutant effect which equals the sum of the simple mutant effects located
within this region, it u~ould be hazardous to consider that smaller de&
ciencies within this region would produce no phenotypic effect. One might


568                  B.tRB:\R,% McCLINTOCK
object that not all the ring chromosomes giving mutant effects were de-
tectibly reduced in chromatin content, for in four of them no deficiency
was detected and in two cases the ring chromosome was enlarged. In all
four cases where no deficiency in the ring chromosome was detected, simple
mutants were produced (brown II, brown III, pink, pale green). If, in
each case, the phenotypic expression is due to a minute deficiency, it is
probable that detection of a single minute deficiency at the meiotic pro-
phase in maize would be extremely difficult. In the cases of the two en-
larged ring chromosomes it has been shown (table 6) that the characters
produced by these two ring chromosomes, brown-pink II and brown-
blotch-dries I, are identical with the characters produced by the reduced
rings brown-pink I and brown-blotch-dries II, respectively. The enlarge-
ment does not mean that there is no deficiency in these ring chromosomes,
for an enlargement may have occurred in an aberrant mitosis subsequent
to an aberrant mitosis which deleted a segment from the ring chromosome.
Following this deletion, no further duplication of the remaining chromatin
could restore the original loss. Neither should subsequent deletions of
duplicated segments cause any change in the expression of the character
produced by such a ring chromosome. It should be emphasized that this
proved to be true in the case of the enlarged brown-pink II ring.
The above considerations are the basis of the inferential evidence that
the plant characters associated with altered ring-shaped chromosomes are
produced as a consequence of homozygous deficiencies. In a previous sec-
tion, the chromatin constitutions of the altered ring chromosomes have
been considered briefly. It was pointed out that the brown-blotch-dries II
ring, the light-green-poor growth ring, and the brown-light green-poor
growth ring were all smaller than the normal four chromomere R-I ring.
In the Df-r/Df-r or Df-r/Df-z plants, the cells which contain only one
such ring must be homozygous deficient for some chromatin within the
limits of the deficiency in the Df-r chromosome. All three of these reduced
ring chromosomes may be distinguished from one another by differences
in thei,r chromomere constitution. Therefore, they do not have exactly the
same deficiencies. Furthermore, each of these three rings produces, respec-
tively, a particular and distinguishable type of compound mutant charac-
ter. It would be difficult to escape the conclusion that the obviously differ-
ent total deficiencies in these three rings are the cause of the obviously
different total group of mutant characters that each of these three ring
chromosomes produces. If one did attempt to account for these observa-
tions on some other bases, it would be necessary to assume that each de-
ficiency in these three cases was without a distinct phenotypic effect-
which hardly seems probable, considering the very striking compound
phenotypic effect that a total deficiency of all four chromomeres produces.


            HO.MOZYGOUS DEFICIENCIES IN M;ZIZE        569
On the basis of the above inferential and observational evidence, it is
concluded that the characters exhibited in tissues containing an altered
ring chromosome are caused by homozygous deficiencies, each character
being caused by a relatively minute but specific deficiency, and that the
compound mutant characters are caused by the removal of two or more
such segments from the genomic complement.

CONCLUSION

This paper presents only a summary of the evidence which has led to
the conclusion that some phenotypic characters in maize may be produced
by homozygous minute deficiencies. A complete account of all the evidence
obtained from altered ring chromosomes-their origin, their gametic trans-
missions both male and female, their stability, the variation in phenotypic
expression of the various mutants when combined with different genotypes,
the cytological analysis of the chromatin constitutions of the various al-
tered ring chromosomes, the appearance of plants with various combina-
tions of three or more altered ring chromosomes, the rate of production of
altered ring chromosomes, evidence showing that the mutants do not re-
vert to normal, and other phenomena related to the behavior of these ring
chromosomes-could not be included in this paper due to limitation of
space. The evidence of particular interest will be reported in separate pub-
lications. It is hoped, however, that sufficient evidence has been included
to indicate the nature of the method of attacking this problem and to in-
dicate the type of evidence obtained. Again, due to limitation of space, an
adequate discussion of the relation of homozygous deficiencies giving mu-
tant effects in maize to similar phenomena in other organisms cannot be
undertaken here. It should be pointed out, however, that the number of
analyzed cases is very limited and is best represented by the yellow mutant
in Drosophila (EPHRUSSI 1934; STERN 1935; MULLER 1935; DEMEREC
1934, 1936; DEMEREC and HOOVER 1936; KAUS 1939).

In conclusion, the author wishes to point out the possible usefulness of
the evidence presented in this paper in formulating a prediction as to the
nature, the location, and the crossover values to be expected of mutants
not as yet obtained in the normal complement of maize. The ring mutants
pink, brown, blotch-dries, and pale green are readily transmitted through
the pollen, although the relative efficiency in competition with grains
carrying an unmodified complement could not be determined. It is possible,
however, that through natural causes or by X-ray or ultra-violet radiation,
these mutants would be produced in a normal rod-chromosome 5. Pale
green and brown should produce viable plants, but pink and blotch-dries
would result in plants which die in the seedling stages. A mutant (brown
mid-rib, bm, JORGENSON 1931), allelic and indistinguishable from the


570                 R.\Kk\R;\ hIc(`I.lNl`O~`K
brown protluc4 by the ring chromosome mutants, has been isolated. r\l-
though it may not be concluded from observational evidence that the bvz
mutant in the normal rod-chromosome j is caused by a homozygous minute
deficiency, it may be stated, however, that a deficiency of the locus of the
dominant allele of bm will reproduce in all details the phenotypic ex-
pression of bm. Pink, blotch-dries, and pale green remain to be isolated.
Following isolation, pink and blotch-dries should prove to be very closel>
linked to the known mutant bm, whereas pale green should show a small
amount of crossing-over.

SUMMARY

It is the purpose of this paper to show that viable mutants in maize ma!
be produced by homozygous minute deficiencies. The aberrant mitotii
behavior of ring-shaped chromosomes has been the method of obtaining
a large number of such deficiencies. These deficiencies are located within
the limits of a relatively small segment of the genomic complement corn-
posed of the proximal 9 chromomeres of the short arm of chromosome 5.

Mutants arise following changes in the chromatin constitution of the
ring-shaped chromosomes. These changes are produced following aberrant
behavior of the ring-shaped chromosome in some of the mitotic divisions.
The types of mutants which arc produced by the altered ring-shaped
chromosomes are simple? composed of a single recognizable character or
compound, composed of two or more of the characters recognizable as
simple mutants. In a number of cases it has been possible to isolate the
altered ring-shaped chromosomes which produce the simple or compountl
mutants.

Through appropriate tests, it has been proven that the compound mu-
tants are the products of two or more of the simple mutants. One group of
mutants has been located within the limits of the proximal 4 chromomeres
of the short arm of chromosome 5. The second group of mutants has been
located within the limits of the next five chromomeres.

Evidence is prese'nted which leads to the conclusion that each mutant
character, whether appearing as a simple mutant or in combination with
other mutants, is produced by a homozygous minute deficiency, each
mutant character being associated with loss of a particular minute seg-
ment. The simple mutants are associated with loss of one such segment;
the compound mutants are associated with loss of two or more such seg-
ments.

                     LITERATURE CITED

I<PHRCSSI, B.. 1934 The absence of autonomy in the development of the effects of certain deticien-
ties in Drosophila melanogavter. Proc. Sat. .kad. Sci. 20: 410-412.
~EblERI:C. .\I.. 1934 Biological action of small deficiencies of S chromosome of Drrxr~plrib~ tueltrm-
  poster. Proc. Sat. Acad. Sci. 20: ,354 -359.


             HOJIOZYGOUS DEFICIENCIES IN MrZIZE          571
  1936 I~requency of "cellLlethals" among lethals obtained at random in the S chromosome of
  Zhsojdzilc~ ~~lamgoder. Proc. Sat. .\cad. Sci. 22: 350-354.
~>LMERFX, M., and 11. I<. HOOVER, 1936 Three related X chromosome deficiencies in Drosophila.
  J. Hered. 27: 206 2 ~2.

JORCES~OK. I,. R., 193~ Brown midrib in maize and its linkage relations. J. Amer. Sot. Agron.
  23: 54T 557.

K.\LISS, S.. 1939 The effect on development of a lethal deficiency in Drosophilrr rrrelorrcjin.sler:
  with a description of the normal embryo at the time of hatching. Genetics 24: 244-270.
~~KLISTOCK. I<., 1933 The association of non-homologous parts of chromosomes in the mid-
  prophase of meiosis in %etr rrt(r~.\. Z. Zellf. mik. Anat. 19: '91-237.
1938 The production of homozygous deficient tissues with mutant characteristics by means
  of the aberrant mitotic behavior of ring-shaped chromosomes. Genetics 23: 3rjm376.

Sltx~:~, H. J., ,935 .1 viable two-gene deficiency. J. Hered. 26: 469-478.
STERN, C.. 19.35 The effect of yellowscute deficiency on somatic cells of Drosophila. Proc. Sat.
  hcad. Sci. 21: 3741379.

S~rno, K. R.. `940 Tnhcritance in .Yirotioncr r&w~r~r. XV. Carmine-white variegation. J. Hered.
  31: `9 24