Trials on Cross and Mutation Breeding of Rice for Adaptability
to Nature Farming
Hirokazu NAKAI*, Yoshiyuki NARA*, Chiharu SUZUKI*, Hiroyasu TABUCHI*,Shigeki NOZAWA** and Yoshihiro HASE*** Institute for Agriculture, Medicine, and the Environment,** Takasaki Advanced Radiation Research Institute, National Institutes for Quantum
and Radiological Science and Technology (QST)
Summary
We have tried to develop rice varieties, through cross and mutation breeding foradaptability to nature farming in these past more than ten years. We succeeded inselecting breeding lines for adaptability to nature farming, in the hybrid populations
from the cross of representative native varieties of Japan, ‘Asahi’ and ‘Kamenoo’. Twobreeding lines, namely AKH2 and AKH4, which were of high yield with high eating
quality in nature farming, were selected from the cross breeding. However, the selected
breeding lines were too late for heading date to be adaptable to wide range of Japanisland other than southern part of Japan e.g. Kyushu district. We tried to improve thenegative trait for late heading of the breeding lines through the use of induced mutations
within five years. This paper refers to the breeding process and discuss on the merit ofmutation breeding and on how to use the technique for gaining adaptability to low input
sustainable agriculture.
Introduction
Conventional agricultural systems dependent on chemical fertilizer and agriculturalchemicals should be converted to low input sustainable agriculture with less amount ofchemicals in the situation of serious climate change covered in the world. It is reported
that more than 20% of all greenhouse gases, e.g. CO 2 , CH 4 , and N 2 O, might be emitted
from the field of agriculture (Nagano et al., 2012, Skinner, 2014). We have tried toperform cross breeding combined with use of induced mutations of rice for adaptability
to nature farming, aiming at developing the system of sustainable agriculture. Nature
farming is defined in this paper as a farming system that uses only residue of rice plants
as the nutrition with no agrochemicals and chemical fertilizer (Okada, 1953). We arereporting a case of the breeding performed in Yunomae-cho, Kumamoto-prefecture,
Japan. The AKH2 breeding line was finally selected for high yield and high eating19quality in the nature farming condition, from the hybrid population derives from thecross between Japanese native varieties, ‘Asahi’ and ‘Kamenoo’. However, the AKH2
breeding line was found to carry weak points for late heading and long culm. We havethereupon tried to improve the demerits of late heading and long culm through use ofinduced mutations. Accordingly, some breeding lines with earlier heading date and/or
shorter culm length as well as higher yield in the low input conditions were selected. The
selected MAKH2 breeding line was noticed as promising new rice variety adaptable tonature farming or low input sustainable agriculture.
This paper refers to the breeding process and discuss on the merit of mutation
breeding and on how to use the technique for gaining adaptability to low input
sustainable agriculture.
Materials and methods
In 2005, Japanese native varieties, Asahi and Kamenoo were crossed at the Ohito
Experimental Farm located in Izunokuni-city, Shizuoka-prefecture located at almost
center of the main island of Japan. The 1,638 F 2 plants derived from 16 F 1 plants weregrown with planting space of 30 cm × 20 cm in the fields of nature farming of Ohito
Experimental Farm in 2007. All the harvested rice straws and 100 kg rice bran wereapplied to the rice fields as nutritional materials. The 140 plants were selected by eyemeasurement from all the F 2 plants in the fields. In 2008, 140 F 3 lines originating fromthe selected F 2 plants were grown and 36 F 3 lines were selected by eye measurement inthe field. The selected F 3 lines were measured for wight of grain and eating quality using
the measuring instrument (Kett Electric Laboratory, Tokyo, Japan).
Table 1 shows the breeding process performed from 2009 to 2017 in Yunome-town,
Kumamoto-prefecture in Kyushu-district, Japan. In 2009, seeds of the selected 36 F 3lines were sent to a farmer, Mr. SHIIBA, Takema in Yunomae-town, Kumamotoprefecture to be grown as F 4 lines in the fields of nature farming. In the selection tests,
any nutritional matters were never applied to the breeding field.
Ten or less breeding lines were used for adaptation and yield tests from 2013 to 2015,
after selection tests up to 2012. In the adaptation and yield tests, the selected 10 or lesslines were grown in randomized block design with three replications (60 plants weregrown per one line of one block). Grain yield and eating quality were measured as wellas main agricultural traits, e.g. culm length, panicle length, number of panicles per plantand so forth. Two breeding lines, AKH2 and AKH4, were finally selected for registeringcommercial cultivar patent. AKH2 was, in fact, registered as commercial cultivar‘Kumaminori’ in 2018.
20

Table 1. The process of cross breeding of native varieties, Asahi and Kamenoo, performedin the farmer’s fields in Yunomae-town, kumamoto-prefecture, Kyushu-district(2009~2017).
Year200920102011201220132014201520162017

F4F5F6F7F8F9F 10F 11F 1236231714107652Hybridgeneration andnumber of lines
grown

However, both breeding lines carry a week point for late heading not to be adaptable
to wider range to the northern part in Japan. In such a situation, we tried to improvethe demerit of the breeding lines through the use of mutation breeding. In 2013, about
5,000 F 8 seeds of AKH2 and AKH4, were irradiated by gamma-rays of 200 Gy at Japan
Atomic Energy Agency (currently Takasaki Advanced Radiation Research Institute,National Institutes for Quantum and Radiological Science and Technology). About 4,000
M 1 plants each of AKH2 and AKH4 were grown in the fields at the Ohito Experimental
Farm. Seed fertility was checked using randomly selected 50 M 1 plants. The selection
process for early heading of the mutants is shown in Table 2. As shown in the Table 2,
after 5 years since the irradiation time, two noticeable mutants were selected from the
population of AKH2 in 2018.
Table 2. Selection process for early heading of rice mutants from the irradiatedpopulations of AKH2 and AKH4.
OriginallineAKH2
AKH4

Year and generation
2013

2014

2015

2016

2017

2018

M1

M2

M3

M4

M5

M6

4,428N*

15,840N

119L**

11L

6L

2L

14,400N

105L

14L

6L

0L

48%***
3,936N
45%****N: number of plants. **L: number of lines.
*** average seed fertility for 50 M 1 plants.

21

Results
Fig.1 shows grain yield and eating quality of the breeding lines, AKH2 and AKH4,
which were selected from cross breeding of Asahi and Kamenoo. In the figure,‘Hinohikari’ was put as a control because of it being representative leading variety inKyushu district. As seen in the figure, grain yield of the AKH2 was higher than AKH4and Hinohikari. As for eating quality, significant difference was not found among the two

700

75.0

650

74.0
73.0

600

72.0

550

71.0

500

70.0

450

69.0
68.0

400

67.0

350
300

eating quality

grain yield（㎏/10a）breeding lines and Hinohikari.

66.0
AKH2

AKH4

grain yield(kg/10a)

Hinohikari

65.0

eating quality

Fig.1. Grain yield and eating quality of the breeding lines, AKH2 and AKH4, incomparison with control (Hinohikari) in the nature farming (from results for adaptationand yield tests in 2014).

We requested some farmers, who were concerned with nature farming in the Kyushu
district, to grow the three lines using farm machines. Fig.2 shows one of the results fromthe farmers on grain yield and eating quality. As seen in the figure, the AKH2 showed
the highest yield. Eating quality of rice was clearly higher in the two breeding lines thanin Hinohikari, suggesting that eating quality is to be higher in the selected breedinglines than in the popular leading variety, Hinohikari in the Kyushu district. We finallyselected the AKH2 breeding line for registering commercial variety in 2018.

226844067420
40066

3806536034064320300

AKH2

AKH4 Hinohikari

AKH22016

AKH4 Hinohikari

Eating quality

Grain yield（kg/10a）

460632017

Grain yield（kg/10a）

Eating quality

Fig. 2. Grain yield and eating quality of breeding lines, AKH2 and AKH4 selected fromthe cross breeding in case of being grown by planting machine by a farmer in Kumamoto
prefecture in 2016 and 2017.
Fig.3 shows heading date and culm length of the breeding lines of AKH2 and AKH4comparing with Ashahi and Kamenoo (parent varieties) and Hinohikari (arepresentative leading variety in Kyushu area). As seen in the figure, heading date of
the both breeding lines was almost same as one of the parent varieties, Asahi, which wassignificantly later in heading date than Kamenoo and Hinohikari. It was noted thatheading date of the selected breeding lines was significantly later than that of the local
leading variety, Hinohikari. It may be inconvenient for farmers in the local area to growthe breeding lines because of it being late in heading date. Culm length of both breedinglines was almost same as that of the parent varieties, Asahi and Kamenoo. However, it
was found that culm length of the breeding lines was significantly longer than that ofHinohikari. We tried to improve the weak point for late heading of the breeding lines to
get wider adaptability to grow in northern part of Japan.
Fig.4 shows the frequency distribution for heading date of M 3 mutant lines
originating from irradiation of gamma-rays to F 8 seeds of the breeding lines of AKH2and AKH4. As seen in the figure, the earliest heading date of the mutant originating
AKH4 was put in 21~31th in July. Heading date of total 15 mutant lines from both thebreeding lines was put in 1 ~10th in September, whereas heading dates of AKH2 andAKH4 were respectively 8th and 6th in September. Heading date of the mutant lines put
in 1~10th in September seen in the figure was earlier than that of both the breedinglines, AKH2 and 4. Anyhow, mutant lines put in 1~10th in September were not so23

practically effective as breeding materials for early heading.

9/18
9/13

100

9/880

9/3
8/29

60

8/24

40

8/19

20
0heading dateculm length (cm)

120

8/14
AKH2

AKH4

asahi

kamenoo hinohikari

8/9culm length (cm)heading dateFig. 3. Heading date and culm length of the breeding lines, AKH2 and AKH4 selectedfrom the cross combination of Asahi and Kamenoo (2014).

12Frequency

1086
4
2
02131July

110Aug.

1120Aug.

2131Aug.

110Sep.

Heading dateFig. 4. Frequency distribution of heading date of the mutant lines originating fromirradiation of gamma-ray of 20 krad to F 8 seeds of AKH4 (white bar in the figure) andAKH2 (black bar in the figure).

24Fig.5 shows culm length and heading date of 6 mutant lines selected as breeding
materials for the relevant breeding purpose, which were originating from irradiation ofgamma-rays to F 8 seeds of AKH2. As seen in the figure, heading date of all the mutantlines selected was significantly earlier than that of control, AKH2. Heading date (12thin August) of the mutant line, MAKH2, was about one month earlier than AKH2. In caseof MAKH2-7, heading date was about one week earlier than the control (AKH2). Culm
length of the mutant lines was significantly lower than that of the control, excepting one
case of MAKH2-7. We, after all, selected two mutant lines, MAKH2-6 and MAKH2-7, ofwhich heading date are earlier than control, AKH2, with same or more grain yield as thecontrol.
Fig.6 shows plant type of the selected M5 mutant lines, MAKH2-7 and MAKH2-6
comparing with control, AKH2. It was found that MAKH2-7 carried much higher numberof panicles comparing with the control, though the culm length was as high as the control.
We have finally selected the MAKH2-7 mutant line as a candidate for registering new10090807060504030
20
100

9/22
9/12
9/2
8/23
8/13
8/3
7/24
7/14

Fig.5. Culm length and heading date of M 4 mutant lines (2017) originating fromirradiation of gamma-rays of 20 krad to F 8 seeds of AKH2.

25

Heading dateCulm length（cm）

rice cultivar for adaptability to nature farming or low input sustainable agriculture.

Fig.6. Plant type of the selected mutant lines, MAKH2-7 and MAKH2-6 and control,
AKH2.

Discussion
We succeeded in development of new rice variety, Kumaminori, through cross ofnative varieties, Asahi and Kamenoo, which shows higher grain yield and better eating
quality in the nature farming or low input sustainable agriculture. The new variety has
been released to farmers to grow in Kyushu district, Japan. However, heading date of
the variety is too late to be adaptable to wider range toward northern part of Japan. It
is noted that farmers are not to be suffering from lodging due to long culm length of thenew variety, because of its strong culm. Farmers are, in general, used to be sufferingfrom lodging of rice in Kyushu district, where is a route of typhoon in autumn season. Itwas practically proofed for strong culm of the new variety, Kumaminori that plants ofthe variety were not fell down when the typhoon came in 2008, while the modern leading
variety, Hinohikari was done (Fig.7).
We have also succeeded in improving late heading date of the breeding line (AKH2)
or new variety, Kumaminori within only five years through use of induced mutations. Itis, in general, to take more than ten years in case of cross breeding for completing
breeding purpose. In fact, we have taken more than 13 years to develop the new variety,Kumaminori.

26

Fig.7. Comparison of scenery of the rice fields of Kumaminori in nature farming andHinohikari in conventional one when attacked by the typhoon in 2018.

The other merit of mutation breeding is to improve a target character without
altering other desirable traits of the original variety. It is supposed that the selectedmutant line, MAKH2-7 is to be inherited with desirable characters from the originalvariety, Kumaminori, e. g. high grain yield and eating quality in the condition of nature
farming or low input sustainable agriculture. It was especially noted that the originalvariety, Kumaminori (AKH2) has been proved from epidemiological survey to carry ahealing function to atopy and allergic disease. It was found earlier by Hiroshi
HASEGAWA that the Yukihikari variety developed in Hokkaido was to carry healing
function to atopy or allergic disease, whereas the representative leading variety,
Kirara397 in Hokkaido tended to cause the disease (Miura et al., 2003; Hasegawa, 2019).
Representative native variety of Japan, Kamenoo and Ahahi were found to carryhealing function to atopy or allergic disease (Sakuma and Komatsu, 2018, The Japan
Agricultural News, 13th December 2019). The varietal difference of rice for ability toreact to the disease was earlier found to be controlled by gene systems (Monnma, 2005;Sonoyama et al., 2009). The results of epidemiological tests suggest that Kumaminori or
the AKH2 breeding line could be inherited healing function to allergic disease from27

mother varieties, Asahi and Kamenoo. We would expect that the MAKH2-7 mutant line
would carry the healing function. Epidemiological tests for allergic disease of the mutantline are in progress. We are now taking a step for registration of the mutant line,
MAKH2-7 for commercial variety.
We have found through the breeding of rice adaptable to nature farming that nativevarieties could be effective materials for breeding of rice for nutritious foods, as well as
for adaptability to low input sustainable agriculture. We also would pay attention to theidea proposed by Indonesia and the Philippines in the sub-project “Mutation Breeding ofrice for Sustainable Agriculture (2013~2017)” that native varieties can be effective
breeding materials for mutation breeding of rice adaptable to low input sustainableagriculture (Nakai, 2018).
In conclusion, we propose that mutation breeding would be useful tool fordevelopment of rice varieties for adaptability to low input sustainable agriculture. Inaddition, we want to suggest that the native varieties be used as effective breeding
materials for mutation breeding of rice for adaptability to low input sustainableagriculture, contributing to rehabilitation of the earth environment and human health.
Acknowledgment
We would thank to Mr. Takema SHIIBA for offering rice fields of the breeding and forkind support on the works. We also would like to give our thanks to Nature Farming andCulture Agency of MOA for financial support for the breeding project. We want to express
our gratitude to Ms. Aki KOIKE of NSRA for kind support on the trials.
References
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