1) Institute of Genetic Resources, Faculty of Agriculture, Kyushu University, Hakozaki, Fukuoka, 812 Japan
2) Plant Breeding and Genetics Research Laboratory, Japan Tabacco Inc., Iwata, Shizuoka, 438 Japan
3) Yamaguchi Women's University, Sakurabatake, Yamaguchi, 753 Japan
Glutelin, a major storage protein in starchy endosperm in rice, is synthesized as a precursor polypeptide of 57 kDa, and cleaved into two subunits of glutelin a (40 kDa polypeptide) and fl (20 kDa polypeptide) by post-translational modification (Yamagata et al. 1982; Takaiwa et al. 1986; Masumura et al. 1989). Recently, Kumamaru et al. (1988) obtained six 57-H mutants with a high content of 57 kDa polypeptide from a Japonica rice cultivar "Kinmaze" by MNU treatment, and demonstrated that one of those mutants, CM 1787, possessed a new recessive gene, esp-2, on chromosome 11 (Kumamaru et al. 1987). In this study, we analysed two other 57-H mutants, EM61 and EM305.
Judging from the intensity of stained bands of both glutelin subunits (a and after SDS-PAGE, EM61 and EM305 were so analogous and were almost the same as Kinmaze in their compositions and contents of glutelins. In contrast, CM1787 showed weakly stained bands compared with those of Kinmaze.
F`1` seeds from a cross between EM61 and Kinmaze showed the mutant phenotype for the 57-H character, and the segregation for normal: mutant types in F`2` seeds fitted well to a 1 :3 ratio, indicating that the 57-H character of EM61 was controlled by a single dominant gene. The F`1` seeds of EM305 and Kinmaze were normal, and the segregation in F'2' seeds fitted a 3 normal: 1 mutant ratio, showing that the 57-H character of EM305 was governed by a single recessive gene.
Table 1 shows the phenotype of F`1` seeds and the segregation pattern in F`2` seeds derived from a diallel cross of CM1787, EM61 and EM305. The F`1` seeds between EM61 and CM1787 (Cross A) showed the 57-H mutant phenotype, and the F`2` seeds were classified into normal, CM1787 and EM61 types. Since the double mutant phenotype was not distinguishable from the CM1787 type, segregation of these three types fitted a 3 :4: 9 ratio. These results suggest that the 57-H mutant gene of EM61 is not allelic to esp-2 of CM1787, and that esp-2 is epistatic to the 57-H mutant gene of EM61.
Between EM305 and CM1787 (Cross B), the F`1` seeds were normal for the 57-H character, and the F`2` seeds were classified into normal, CM1787 and EM 305 types. As the double recessive type was not distinguishable from the 57-H mutant type of CM1787, the F`2` ratio fitted 9:4:3, indicating that the 57-H mutant gene of EM305 is not allelic to esp-2 and that esp-2 is epistatic to the 57-H mutant gene of EM305.
Table 1. Segregation pattern of 57-H character in F2 seeds of crosses between
three 57-H mutants, EM 61, EM 305 and CM 1787
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Cross Combination F`1` seed F`2` segregation Chi2
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Normal CM1787 EM61 Total (3:4:9)
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A EM61 x CM1787 57-H 19 24 57 100 0.05
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Normal CM1787 EM305 Total (9:4:3)
B EM305xCM1787 Normal 52 26 22 100 0.92
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Normal EM61 + EM305 Total
c EM61 x EM305 57-H 13 87 100
12 88 100
Total 25 175 200
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The F`1` seeds between EM61 and EM305 (Croos C) showed the mutant phenotype,
and the F`2` seeds were classified into normal and 57-H types as the parents
were similar and indistinguishable in their gel profiles. The occurrence of
normal type in the F`2` seed, however, suggests that genes controlling the 57-H
character in the parents are located at different loci.Furthermore, trisomic analysis of the two 57-H mutants, EM61 and EM305, showed that in crosses between each mutant and triplo 9 plants, the F`2` segregation pattern fitted the trisomic ratio (Tables 2 and 3). This indicates that the two mutant genes, one is EM61 and dominant, and the other is EM305 and recessive, are both located on chromosome 9 and linked. These genes are designated as Gup-1(t) and gup-2(t), respectively, implying glutelin-precursor mutant. Their recombination value was estimated to be 0.2925 from the data in Table 1 (Cross C).
Table 2. Segregation of normal and 57-H in F`2` seeds set on the trisomic
F`1` plants derived from the crosses between seven types of
trisomics and EM61
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Type of Trisomics F2 segregation Total Chi2
Normal 57-H (1:3)
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Triplo 9 89 111 200 40.56**
Triplo 4 20 60 80 0.00
Triplo 5 21 59 80 0.07
Triplo 6 23 57 80 0.60
Triplo 8 22 58 80 0.27
Triplo 11 21 59 80 0.07
Triplo 12 15 64 79 1.52
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**Significant at 1% level
Table 3. Segregation of normal and 57-H in F`2` seeds set on the trisomic
F`1` plants derived from the crosses between nine types of trisomics
and EM 305
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F2 segregation Chi2
Type of Trisomics Total
Normal 57-H (3:1)
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Triplo 9 291 26 317 47.71**
Triplo 4 28 10 38 0.06
Triplo 5 61 19 80 0.07
Triplo 6 28 10 38 0.06
Triplo 7 87 35 122 0.89
Triplo 8 97 22 119 2.69
Triplo 10 64 16 80 1.07
Triplo 11 32 8 40 0.53
Triplo 12 29 9 38 0.04
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ReferencesKumamaru, T., H. Satoh, N. Iwata, T. Omura and M. Ogawa, 1987. Mutant for rice storage proteins. III. Genetic analysis of mutants for storage proteins of protein bodies in the starchy endosperm. Jpn. J. Genet. 62: 333-339.
Kumamaru, T., H. Satoh, N. Iwata, T. Omura, M. Ogawa and K. Tanaka, 1988. Mutants for rice storage proteins. 1. Screening of mutants for storage proteins of protein bodies in the starchy endosperm. Theor. Appl. Genet. 76: 11-16.
Masumura, T., K. Kidzu, Y. Sugiyama, N. Mitsukawa, T. Hibino, K. Tanaka and S. Fujii, 1989. Nucleotide sequence of cDNA encoding major rice glutelin. Plant Mol. Biol. 12: 723-725.
Takaiwa, F., S. Kikuchi and K. Oono, 1986. The structure of rice storage protein glutelin precursor deduced from cDNA. FEBS Lett. 206: 33-35.
Yamagata, H., T. Sugimoto, K. Tanaka and Z. Kasai, 1982. Biosynthesis of storage proteins in developing rice seeds. Plant Physiol. 70: 1094-1100.