Identification and Molecular Analysis of Four New Alleles at the W1 Locus Associated with Flower Color in Soybean

Abstract

In soybean, flavonoid 3′5′-hydroxylase (F3′5′H) and dihydroflavonol-4-reductase (DFR) play a crucial role in the production of anthocyanin pigments. Loss-of-function of the W1 locus, which encodes the former, or W3 and W4, which encode the latter, always produces white flowers. In this study, we searched for new genetic components responsible for the production of white flowers in soybean and isolated four white-flowered mutant lines, i.e., two Glycine soja accessions (CW12700 and CW13381) and two EMS-induced mutants of Glycine max (PE1837 and PE636). F3′5′H expression in CW12700, PE1837, and PE636 was normal, whereas that in CW13381 was aberrant and missing the third exon. Sequence analysis of F3′5′H of CW13381 revealed the presence of an indel (~90-bp AT-repeat) in the second intron. In addition, the F3′5′H of CW12700, PE1837, and PE636 harbored unique single-nucleotide substitutions. The single nucleotide polymorphisms resulted in substitutions of amino acid residues located in or near the SRS4 domain of F3′5′H, which is essential for substrate recognition. 3D structure modeling of F3′5′H indicated that the substitutions could interfere with an interaction between the substrate and heme group and compromise the conformation of the active site of F3′5′H. Recombination analysis revealed a tight correlation between all of the mutant alleles at the W1 locus and white flower color. On the basis of the characterization of the new mutant alleles, we discussed the biological implications of F3′5′H and DFR in the determination of flower colors in soybean.

Nonsense Mutation Inside Anthocyanidin Synthase Gene Controls Pigmentation in Yellow Raspberry (Rubus idaeus L.)

“…Anthocyanins and carotenoids are considered to be the main pigments involved in coloration of raspberries (de Ancos et al, 1999; Carvalho et al, 2013b). Anthocyanins are water soluble polyphenolic pigments responsible for the colors of many flowers (Chung et al, 2010; Luo et al, 2015; Sundaramoorthy et al, 2016), fruits (Saito et al, 1999; Debes et al, 2011; Liu et al, 2013; Ben-Simhon et al, 2015), and other plant tissues (Gould et al, 2000; Kim et al, 2004; Zhou et al, 2010). Their biochemical role in plants is not fully understood, however, they have been considered to protect tissues from biotic and abiotic stresses, delay in senescence, assist in photosynthetic machinery, delay over-ripening and increase shelf-life of fruits and act as scavengers of reactive oxygen intermediates (Zhang et al, 2013, 2016; Landi et al, 2015; Yousuf et al, 2016).…”

Molecular Genetic Analysis of a Novel Recessive White Flower Gene in Wild Soybean

“…Moreover, our phylogenetic analysis of soybean DFR2 genes indicated that the reported kw4 DFR2 sequence itself was questionable because its structure appeared to be either an artificial chimera or natural hybrid between G. soja and G. max. Concern about the significance of the 367-bp deletion in the third intron of the sequence was also expressed by Sundaramoorthy et al (2016).…”

“…When compared with those of Hwangkeum and Clark bearing purple flowers, >20 SNPs and insertion-deletion events were observed in the five intron regions. However, only one synonymous SNP was identified in the exon region, indicating that the same amino acid sequence was encoded by DFR2 in both YWS415 and W4 soybean accessions in the absence of the transposon insertion (Xu et al, 2010;Yan et al, 2014;Sundaramoorthy et al, 2016).…”

“…However, only one synonymous SNP was identified in the exon region, indicating that the same amino acid sequence was encoded by DFR2 in both YWS415 and W4 soybean accessions in the absence of the transposon insertion (Xu et al, 2010;Yan et al, 2014;Sundaramoorthy et al, 2016). KY744345) of YWS415 covered the postulated full-length coding sequence of DFR2.…”

“…Linkage analysis of the markers was performed using CarthaGene version 1.2 (Schiex and Gaspin, 1997). Six fulllength nucleotide sequences of DFR2 (Xu et al, 2010;Yan et al, 2014;Sundaramoorthy et al, 2016) were downloaded for comparison with our two DFR2 sequences. Relationships among the DFR2 sequences were assessed using ClustalW and the neighbor-joining and bootstrap method implemented in MEGA6 (Tamura et al, 2013).…”