ABSTRACT
Time to flowering and maturity in soybean is controlled by loci E1 to E5, and E7 to E9. These loci were assigned to molecular linkage groups (MLGs) except for E5. This study was conducted to map the E5 locus using F2 populations expected to segregate for E5. F2 populations were subjected to quantitative trait locus (QTL) analysis for days to flowering (DF) and maturity (DM). In Harosoy-E5 × Clark-e2 population, QTLs for DF and DM were found at a similar position with E2. In Harosoy × Clark-e2E5 population, QTLs for DF and DM were found in MLG D1a and B1, respectively. In Harosoy-E5Dt2 × Clark-e2 population, a QTL for DF was found in MLG B1. Thus, results from these populations were not fully consistent, and no candidate QTL for E5 was found. In Harosoy × PI 80837 population, from which E5 was originally identified, QTLs corresponding to E1 and E3 were found, but none for E5 existed. Harosoy and PI 80837 had the e2-ns allele whereas Harosoy-E5 had the E2-dl allele. The E2-dl allele of Harosoy-E5 may have been generated by outcrossing and may be responsible for the lateness of Harosoy-E5. We conclude that a unique E5 gene may not exist.
ABSTRACT
BACKGROUND: Flavonol glycosides (FGs) are major components of soybean leaves and there are substantial differences in FG composition among genotypes. The first objective of this study was to identify genes responsible for FG biosynthesis and to locate them in the soybean genome. The second objective was to clone the candidate genes and to verify their function. Recombinant inbred lines (RILs) were developed from a cross between cultivars Nezumisaya and Harosoy. RESULTS: HPLC comparison with authentic samples suggested that FGs having glucose at the 2â³-position of glucose or galactose that is bound to the 3-position of kaempferol were present in Nezumisaya, whereas FGs of Harosoy were devoid of 2â³-glucose. Conversely, FGs having glucose at the 6â³-position of glucose or galactose that is bound to the 3-position of kaempferol were present in Harosoy, whereas these FGs were absent in Nezumisaya. Genetic analysis suggested that two genes control the pattern of attachment of these sugar moieties in FGs. One of the genes may be responsible for attachment of glucose to the 2â³-position, probably encoding for a flavonol 3-O-glucoside/galactoside (1 â 2) glucosyltransferase. Nezumisaya may have a dominant whereas Harosoy may have a recessive allele of the gene. Based on SSR analysis, linkage mapping and genome database survey, we cloned a candidate gene designated as GmF3G2â³Gt in the molecular linkage group C2 (chromosome 6). The open reading frame of GmF3G2â³Gt is 1380 bp long encoding 459 amino acids with four amino acid substitutions among the cultivars. The GmF3G2â³Gt recombinant protein converted kaempferol 3-O-glucoside to kaempferol 3-O-sophoroside. GmF3G2â³Gt of Nezumisaya showed a broad activity for kaempferol/quercetin 3-O-glucoside/galactoside derivatives but it did not glucosylate kaempferol 3-O-rhamnosyl-(1 â 4)-[rhamnosyl-(1 â 6)-glucoside] and 3-O-rhamnosyl-(1 â 4)-[glucosyl-(1 â 6)-glucoside]. CONCLUSION: GmF3G2â³Gt encodes a flavonol 3-O-glucoside/galactoside (1 â 2) glucosyltransferase and corresponds to the Fg3 gene. GmF3G2â³Gt was designated as UGT79B30 by the UGT Nomenclature Committee. Based on substrate specificity of GmF3G2â³Gt, 2â³-glucosylation of flavonol 3-O-glycoside may be irreconcilable with 4â³-glycosylation in soybean leaves.
Subject(s)
Chromosome Mapping/methods , Cloning, Molecular/methods , Flavonols/metabolism , Genes, Plant , Glucosyltransferases/genetics , Glycine max/genetics , Glycosides/metabolism , Amino Acid Sequence , Chromatography, High Pressure Liquid , Flavonols/analysis , Flavonols/chemistry , Gene Expression Regulation, Plant , Glucosyltransferases/chemistry , Glucosyltransferases/metabolism , Glycosides/analysis , Glycosides/chemistry , Inbreeding , Inheritance Patterns/genetics , Molecular Sequence Data , Phylogeny , Polymorphism, Genetic , Recombination, Genetic/genetics , Substrate SpecificityABSTRACT
Soybean (Glycine max) is a major crop in the world. Three new flavonol 3-O-glycosides, kaempferol 3-O-alpha-L-rhamnopyranosyl-(1 --> 4)-[alpha-L-rhamnopyranosyl-(1 --> 6)-beta-D-galactopyranoside] (1), kaempferol 3-O-alpha-L-rhamnopyranosyl-(1 --> 4)-[beta-D-glucopyranosyl-(1 --> 6)-beta-D-galactopyranoside] (4) and quercetin 3-O-beta-D-glucopyranosyl-(1--> 2)-[alpha-L-rhamnopyranosyl-(1 --> 6)-beta-galactopyranoside] (5) were isolated from the leaves of soybean cultivars, together with three known compounds, kaempferol 3-O-beta-D-glucopyranosyl-(1 --> 2)-[alpha-L-rhamnopyranosyl-(1--> 6)-beta-D-galactopyranoside] (2), kaempferol 3-O-beta-D-glucopyranosyl-(1 --> 2)-[alpha-L-rhamnopyranosyl-(1 --> 6)-beta-D-glucopyranoside] (3) and quercetin 3-O-beta-D-glucopyranosyl-(1 --> 2)-[alpha-L-rhamnopyranosyl-(1 --> 6)-beta-D-glucopyranoside] (6), and also common flavonoids. The isolated compounds possess similar structures and high water solubility, and so it was hard to isolate them (in particular 5 and 6) with a normal preparative HPLC system. Their final purification was achieved by a preparative HPLC system equipped with a recycle device.