Identification of Gene-Allele System Conferring Alkali-Tolerance at Seedling Stage in Northeast China Soybean Germplasm.

Salinization of cultivated soils may result in either high salt levels or alkaline conditions, both of which stress crops and reduce performance. We sampled genotypes included in the Northeast China soybean germplasm population (NECSGP) to identify possible genes that affect tolerance to alkaline soil conditions. In this study, 361 soybean accessions collected in Northeast China were tested under 220 mM NaHCO3:Na2CO3 = 9:1 (pH = 9.8) to evaluate the alkali-tolerance (ATI) at the seedling stage in Mudanjiang, Heilongjiang, China. The restricted two-stage multi-locus model genome-wide association study (RTM-GWAS) with gene-allele sequences as markers (6503 GASMs) based on simplified genome resequencing (RAD-sequencing) was accomplished. From this analysis, 132 main effect candidate genes with 359 alleles and 35 Gene × Environment genes with 103 alleles were identified, explaining 90.93% and 2.80% of the seedling alkali-tolerance phenotypic variation, respectively. Genetic variability of ATI in NECSGP was observed primarily within subpopulations, especially in ecoregion B, from which 80% of ATI-tolerant accessions were screened out. The biological functions of 132 candidate genes were classified into eight functional categories (defense response, substance transport, regulation, metabolism-related, substance synthesis, biological process, plant development, and unknown function). From the ATI gene-allele system, six key genes-alleles were identified as starting points for further study on understanding the ATI gene network.

An Improved Genome-Wide Association Procedure Explores Gene-Allele Constitutions and Evolutionary Drives of Growth Period Traits in the Global Soybean Germplasm Population.

In soybeans (Glycine max (L.) Merr.), their growth periods, DSF (days of sowing-to-flowering), and DFM (days of flowering-to-maturity) are determined by their required accumulative day-length (ADL) and active temperature (AAT). A sample of 354 soybean varieties from five world eco-regions was tested in four seasons in Nanjing, China. The ADL and AAT of DSF and DFM were calculated from daily day-lengths and temperatures provided by the Nanjing Meteorological Bureau. The improved restricted two-stage multi-locus genome-wide association study using gene-allele sequences as markers (coded GASM-RTM-GWAS) was performed. (i) For DSF and its related ADLDSF and AATDSF, 130-141 genes with 384-406 alleles were explored, and for DFM and its related ADLDFM and AATDFM, 124-135 genes with 362-384 alleles were explored, in a total of six gene-allele systems. DSF shared more ADL and AAT contributions than DFM. (ii) Comparisons between the eco-region gene-allele submatrices indicated that the genetic adaptation from the origin to the geographic sub-regions was characterized by allele emergence (mutation), while genetic expansion from primary maturity group (MG)-sets to early/late MG-sets featured allele exclusion (selection) without allele emergence in addition to inheritance (migration). (iii) Optimal crosses with transgressive segregations in both directions were predicted and recommended for breeding purposes, indicating that allele recombination in soybean is an important evolutionary drive. (iv) Genes of the six traits were mostly trait-specific involved in four categories of 10 groups of biological functions. GASM-RTM-GWAS showed potential in detecting directly causal genes with their alleles, identifying differential trait evolutionary drives, predicting recombination breeding potentials, and revealing population gene networks.

Genotyping of Soybean Cultivars With Medium-Density Array Reveals the Population Structure and QTNs Underlying Maturity and Seed Traits.

Soybean was domesticated about 5,000 to 6,000 years ago in China. Although genotyping technologies such as genotyping by sequencing (GBS) and high-density array are available, it is convenient and economical to genotype cultivars or populations using medium-density SNP array in genetic study as well as in molecular breeding. In this study, 235 cultivars, collected from China, Japan, USA, Canada and some other countries, were genotyped using SoySNP8k iSelect BeadChip with 7,189 single nucleotide polymorphisms (SNPs). In total, 4,471 polymorphic SNP markers were used to analyze population structure and perform genome-wide association study (GWAS). The most likely K value was 7, indicating this population can be divided into 7 subpopulations, which is well in accordance with the geographic origins of cultivars or accession studied. The LD decay rate was estimated at 184 kb, where r2 dropped to half of its maximum value (0.205). GWAS using FarmCPU detected a stable quantitative trait nucleotide (QTN) for hilum color and seed color, which is consistent with the known loci or genes. Although no universal QTNs for flowering time and maturity were identified across all environments, a total of 30 consistent QTNs were detected for flowering time (R1) or maturity (R7 and R8) on 16 chromosomes, most of them were corresponding to known E1 to E4 genes or QTL region reported in SoyBase (soybase.org). Of 16 consistent QTNs for protein and oil contents, 11 QTNs were detected having antagonistic effects on protein and oil content, while 4 QTNs soly for oil content, and one QTN soly for protein content. The information gained in this study demonstrated that the usefulness of the medium-density SNP array in genotyping for genetic study and molecular breeding.

Allelic variations at four major maturity E genes and transcriptional abundance of the E1 gene are associated with flowering time and maturity of soybean cultivars.

The time to flowering and maturity are ecologically and agronomically important traits for soybean landrace and cultivar adaptation. As a typical short-day crop, long day conditions in the high-latitude regions require soybean cultivars with photoperiod insensitivity that can mature before frost. Although the molecular basis of four major E loci (E1 to E4) have been deciphered, it is not quite clear whether, or to what degree, genetic variation and the expression level of the four E genes are associated with the time to flowering and maturity of soybean cultivars. In this study, we genotyped 180 cultivars at E1 to E4 genes, meanwhile, the time to flowering and maturity of those cultivars were investigated at six geographic locations in China from 2011 to 2012 and further confirmed in 2013. The percentages of recessive alleles at E1, E2, E3 and E4 loci were 38.34%, 84.45%, 36.33%, and 7.20%, respectively. Statistical analysis showed that allelic variations at each of four loci had a significant effect on flowering time as well as maturity. We classified the 180 cultivars into eight genotypic groups based on allelic variations of the four major E loci. The genetic group of e1-nf representing dysfunctional alleles at the E1 locus flowered earliest in all the geographic locations. In contrast, cultivars in the E1E2E3E4 group originated from the southern areas flowered very late or did not flower before frost at high latitude locations. The transcriptional abundance of functional E1 gene was significantly associated with flowering time. However, the ranges of time to flowering and maturity were quite large within some genotypic groups, implying the presence of some other unknown genetic factors that are involved in control of flowering time or maturity. Known genes (e.g. E3 and E4) and other unknown factors may function, at least partially, through regulation of the expression of the E1 gene.