DNA methylation affected by male sterile cytoplasm in rice ( Oryza sativa L.)

Male sterile cytoplasm plays an important role in hybrid rice, and cytoplasmic effects are sufficiently documented. However, no reports are available on DNA methylation affected by male sterile cytoplasm in hybrid rice. We used a methylation-sensitive amplified polymorphism technique to characterize DNA methylation in four male sterile cytoplasms that are widely commercialized in China. In total, 12 pairs of selective primers in combinations of Eco RI and Msp I/ Hpa II amplified 350 bands among four male sterile (A) lines and the corresponding maintainer (B) lines. Sites b1 and b3 were fully methylated only in all the B lines, while b2 was fully methylated only in all the A lines. These results implied a relationship of DNA methylation at these sites specifically with male sterile cytoplasms, as well as male sterility, since the only difference between the A and B lines was the cytoplasm. The DNA methylation was markedly affected by male sterile cytoplasms. WA-type and Yinshui-type cytoplasms affected the methylation to a much greater degree than G-type and D-type cytoplasms, as indicated by the number and degree of methylated sites, ratio of methylated sites, number of fully methylated sites, ratio of fully methylated sites, and polymorphism between A and B lines for these cytoplasms. The genetic distance between the cytoplasm and nucleus for the WA-type is much greater than for G- and D-types because the former is between wild and cultivated species and the latter is within indica subspecies between African and Asian cultivars. This difference in genetic distance may be responsible for the variation in methylation which we observed.