Genetic analysis and molecular characterization of the W4 locus and the k2 Mdh1-n y20 chromosomal region in soybean [Glycine max (L) Merr

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The W4 locus is an important locus to the anthocyanin biosynthesis in soybean flowers and hypocotyls. Mutations at this locus would cause less-pigmented flowers and hypocotyls, such as near-white flowers conditioned by the w4 allele. Its mutable allele (w4m) was suspected to contain an autonomous transposon. In this study, we reported a mutation with pale flowers and green hypocotyls conditioned by a new allele at the W4 locus, w4-p. The dominance of these four alleles is: W4 > w4-m > w4-p > w4. The inbred mutant line (w4p w4-p) was assigned Genetic Type Collection number T369. The W4 locus was positioned on the MLG D2 between Satt386 and nearby telomere with 2.3 cM genetic distance from Satt386. The major components of anthocyanins in soybean flowers were found to be delphinidin and its derivatives, which were reduced in less pigmented flower petals caused by mutations at the W4 locus. Results in this study showed that the pigment reduction was due to low transcript level of the DFR 2 (dihydroflavonol reductase 2) gene. The results also indicated that the W4 gene very likely encoded the DFR2 protein. Three closely linked genes k2, Mdhl-n, and y20 form an unstable chromosomal region. The Mdhl-n mutation in T261 and all the Mdhl-n y20 mutations were suggested to be due to genomic deletions. Four more new mutants in this region were reported in this study. One had tan-saddle seed coat (k2) isolated from a bulk harvest of cv. Kenwood. Three independent mutants (k2 Mdhl-n y20) were reported in the instability experiments. The unstable region was located on MLG H near SSR markers Satt253, Satt279, and Satt314. In addition, an extra putative deletion that covered Satt253, Satt279, and Satt314 was identified in T261. Mapping results indicated the two deletions were separate. An Mdhl contig with size around 96 kb was constructed. Three copies of truncated Calypso5-l like retroelements were found downstream of the Mdhl gene (AF180335), and clustered in a -16 kb chromosomal region. The deletions in all the Mdhl-n or Mdhl-n y20 mutants examined were larger that ~70 kb, but the breakpoints couldn't be identified.