Identification of Genes Responsible for the Synthesis of Glycitein Isoflavones in Soybean Seeds.

Soybean ( (L.) Merrill) isoflavones are among the most important secondary metabolites, with functional benefits for human health. Soybeans accumulate three aglycone forms of isoflavones: genistein, daidzein, and glycitein. Soybean landrace Kumachi-1 does not accumulate malonylglycitin at all. Gene structure analysis indicated that () of Kumachi-1 has a 3.8-kbp insertion, resulting in a truncated flavonoid 6-hydroxylase () sequence compared to the wild-type sequence in Fukuyutaka. Mapping experiments using a mutant line (MUT1246) with a phenotype similar to that of Kumachi-1, with a single-nucleotide polymorphism (SNP) in , revealed co-segregation of this mutation and the absence of glycitein isoflavones. We also identified a mutant line (K01) that exhibited a change in the HPLC retention time of glycitein isoflavones, accumulating glycoside and malonylglycoside forms of 6-hydroxydaidzein. K01 contains an SNP that produces a premature stop codon in (), a novel soybean isoflavone O-methyltransferase () gene. We further analyzed transgenic hairy roots of soybeans expressing () and (). Those overexpressing accumulated malonylglycoside forms of 6-hydroxydaidzein (M_6HD), and co-expression of and increased the level of malonylglycitin but not of M_6HD. These results indicate that and are responsible for glycitein biosynthesis in soybean seed hypocotyl.