Soybean locus qDTF-7 as an example of genetic heterogeneity associated with flowering and maturity time

Genome-wide association studies (GWAS) have become a standard approach for identifying quantitative trait loci associated with diverse phenotypic traits. Further investigation of the locus - specifically, the search for the causal gene and mutation - may present various challenges. One of the challenges is genetic heterogeneity (or locus heterogeneity), when alleles from different closely located genes can influence the same trait. Recently, using GWAS, we found the qDTF-7 locus on soybean chromosome 3, which is associated with flowering time under Novosibirsk conditions. Initially, we identified GmTOE1, an ortholog of TOE1 (TARGET OF EAT1), a known flowering-time regulator in Arabidopsis, as the most likely candidate gene for this locus. Four major haplotypes were identified in GmTOE1, which are associated with soybean flowering and maturity and are likely to provide soybean adaptation to northern latitudes. However, this gene showed only a very weak association with soybean flowering in the Novosibirsk region compared to the Oryol region, suggesting the presence of another gene within the locus that influences flowering time. We therefore re-analyzed genes in the qDTF-7 locus and identified GmRVE8c, an Arabidopsis RVE8 (REVEILLE 8) ortholog, located ~21 kb upstream of GmTOE1; RVE8 is a circadian clock component involved in plant development. After studying the natural variation of the GmRVE8c genes, we found four major haplotypes that arose due to three nonsynonymous substitutions and one 19-bp deletion leading to a frameshift. To identify three haplotypes, GmRVE8chap1, 3, 4, which are predominant in improved soybean cultivars, we developed DNA markers. Using these markers, we genotyped 129 soybean accessions, the developmental time of which had been studied in the Novosibirsk and Oryol regions. Using our data and data from SoyOmics, we found the GmRVE8chap3 and GmRVE8chap4 haplotypes to be associated with late flowering and maturity in soybean. The early-maturing haplotype GmRVE8chap1 is predominant in cultivars from northern regions and is likely associated with the adaptation of soybean to high latitudes. The GmRVE8chap4 haplotype is in complete linkage with the early-maturing allele GmTOE1C, whereas the GmRVE8chap3 haplotype shows strong linkage with the late maturing allele GmTOE1T. Furthermore, the ANOVA results indicate an interaction between GmRVE8c and E1, the major regulator of flowering in soybean. This interaction is manifested as a stronger effect of the GmRVE8chap3,4 haplotypes on flowering and maturity in the genetic background of the e1-as allele compared with E1. Together, these findings define a complex and intriguing locus, which may serve as a possible example of a genetically heterogeneous locus.