Variation in proline-rich proteins and DNA methylation patterns in soybean seed color isolines

The I locus controls pattern expression of anthocyanin pigments in soybean seed coats. Using isolines of soybean cultivars varying only at the I locus, a seed coat cell wall protein that is affected by seed coat color was identified. A cDNA library was constructed using messenger RNA isolated from immature soybean seed coats. A 993 base pair clone corresponding to the protein was isolated and sequenced. The protein was identified as PRP1. It was abundant in immature seed coats of yellow seeded Richland but present at a much reduced level in the black seeded isoline T157. PRP1 was isolated from soybean seed coats using insoluble polyvinylpolypyrrolidone column chromatography. The expression of a similar seed coat protein, PRP2, was not affected by seed color genotype. Thus, a mutation in the anthocyanin pathway appears to affect a specific cell wall protein, PRP1.PRP1 was also affected by the Im locus. Saline soluble proteins were isolated from the seed coats of soybean isolines with the genotypes Im/Im or im/im. PRP1 was found to have decreased in apparent molecular weight in Im lines compared to protein from im genotypes. The shift was also evident in PRP2 extracted from both the hypocotyl hood and mature seed coats.The role of transposable elements in the generation of the mutant T157 was investigated on Southern blots. A methyl sensitive HpaII restriction fragment length polymorphism (RFLP) was evident when genomic digests of Richland and T157 were probed with an internal HindIII fragment from the soybean transposable element Tgm1. RFLPs were also apparent when genomic DNA was digested with HpaII/KpnI. Segregation of seed coat color in the F2 generation of crosses between Richland and T157 did not correspond with restriction fragment length polymorphisms. In five different F2 lines, new band patterns were evident on Southern blots. In one of these lines, the new band pattern was stable through five generations of selfing. The new band patterns were generated by changes in methylation of HpaII sites near Tgm1 related sequences.U of I OnlyETDs are only available to UIUC Users without author permissio