Genetic Analysis And Molecular Mapping Of Five Yellow-Green Leaf Mutants And A Male Sterile Mutant In Rice

The sources of rice mutants are very extensive, which are mainly divided into spontaneous mutation and artificial mutation, and they are also the main ways to select or create new species. The objective of this study is to conduct genetic analysis and molecular mapping of six rice mutants. D77, D83, D86, D91 and yg198 are yellow-green leaf mutants, and they were isolated by EMS mutagenesis. Male sterile mutant 802A was discovered in the F8 population from the crosses between two indica lines, i.e. shuihui 527 and shuhui 881. The results are as follows:The whole plants of D77, D83, D86 and D91 exhibit yellow-green character throughout the growing period. They have similar phenotypes and primary agronomic traits, and the differences of their photo synthetic pigment contents are not significient. Compared with their wild-type parent 10079, the average contents of chlorophyll and carotenoid decreased by 47.1% and 40.4%; the plant height, the number of spikelets per panicle and the 1000-grain weight reduced by 6.7%,8.9% and 10.2%, respectively; the flag leaf width increased by 13.3%. The result of electron microscopic observation revealed that the chloroplasts in the D77 mutant were out-of-shape, and some chloroplasts were twisted. Grana stacks arranged loosely and appeared to be less dense compared to those of the wild type. Genetic analysis showed that all of the yellow-green leaf characters of D77, D83, D86 and D91 were controlled by one pair of recessive nuclear genes. Genetic mapping of the mutant genes were conducted by using molecular markers and F2 mapping populations of D77/Zhefu802,D83/Zhefu802,D86/Zhefu802 and D9//Zhefu802. The results suggested that all of the mutant genes of D77, D83, D86 and D91 were mapped on the short arm of rice chromosome 2, nearby SSR marker RM110, between InDel markers Ch2-27 and Ch2-32, with average genetic distances of 1.5 and 1.6 cM. The crosses among D77, D83, D86 and D91 showed that the seedling leaf colors of all F1 plants were similar to that of their parents, and all of they were yellow-green. Therefor, we can say that the mutant genes of D 77, D83, D86 and D91 are alleles, and they are considered to be a new yellow-green leaf gene in rice. The yellow-green leaf mutant yg198 was isolated by chemistry mutagenesis, and its whole plant exhibited yellow-green character throughout the growing period, too. Compared with its wild-type parent 10079, the contents of chlorophyll and carotenoid decreased by 45.3% and 45.6%; and at the maturity, the number of productive panicles per plant, seed setting rate and plant height reduced by 14.4%,10.7% and 7.4%, respectively. The result of electron microscopic observation revealed that the chloroplasts in the yg198 mutant were out-of-shape. A lot of cystic structures and poor thylakoids were observed in the chloroplasts of the yg198 mutant, and grana stacks appeared to be less dense compared to those of the wild type. Genetic analysis showed that the yellow-green leaf character of the yg198 mutant was controlled by one pair of recessive nuclear genes. Genetic mapping of the mutant gene was conducted by using 771 yellow-green leaf individuals from the F2 mapping population of yg198/Zhefu802. Finally, the mutant gene was mapped between InDel markers 13 and 14 on the long arm of chromosome 3, with genetic distances of 0.07 and 0.19 cM, respectively, and with physical distance of 44.2 kb, in which eight predicted genes had been annotated. Sequencing analysis of these candidate genes between the mutant and its wild-type revealed the single base change (G1 522A) of the gene for magnesium-chelatase Ch1D subunit resulted in a missense mutation (A508T) in the encoded product. The same gene mutation caused by OsCh1D (Chlorina-1) was documented previously. The Chlorina-1 mutant displays a severe yellowish-green leaf phenotype only at the seedling stage, and the abnormal leaf color is first observed on the leaves of 2-to 3-week-old seedlings, while the yg198 mutant exhibits yellow-green character throughout the growing period. The different phenotypes of the two mutants may be caused by the different mutational sites of OsCh1D genomic sequence.The male sterile mutant 802A was discovered in the Fg population from the crosses between two indica lines, i.e. shuihui 527 and shuhui 881. The anther of the 802A mutant was slender and milky-yellow, and remained unbroken after flowering. Its pollen primarily belonged to typical abortion type. Meanwhile, the mutant showed that the glumes were thinned and twisted; the flag leaves were narrowed, shortened and frizzled; the basal part of panicles were enclosed in the sheath. The previous research results in our lab indicated that the male sterility of the 802A mutant was controlled by a pair of recessive nuclear genes, and the mutant gene was mapped between SSR markers RM6266 and RM3513 on the long arm of chromosome 3, with genetic distances of 3.9 and 0.6 cM, respectively. In order to further map the targeted gene, we developed five polymorphic InDel markers:S1, S2, S3, S4 and S5, and finally, the gene was found between InDel markers S2 and S5. The genetic distances from the target gene to the two InDel markers were 0.6 and 0.3 cM, respectively. At the same time, the mutant gene co-segregated with InDel markers S3 and S4 in 167 F2 male sterile individuals from the mapping population. S2 and S5 markers mapped a 244-kb DNA region on three bacterial artificial chromosomes, AC147427, AC145383 and AL713930. Compared with known male sterile genes, the mutant gene of 802A was considered to be a novel recessive male sterile gene in rice, and designated tentatively as ms92(t).