Soybean is an important oil seed crop, but very few high-density genetic maps have been published for this species. Specific length amplified fragment sequencing (SLAF-seq) is a recently developed high-resolution strategy for large scale de novo discovery and genotyping of single nucleotide polymorphisms. SLAF-seq was employed in this study to obtain sufficient markers to construct a high-density genetic map for soybean. In total, 33.10 Gb of data containing 171,001,333 paired-end reads were obtained after preprocessing. The average sequencing depth was 42.29 in the Dongnong594, 56.63 in the Charleston, and 3.92 in each progeny. In total, 164,197 high-quality SLAFs were detected, of which 12,577 SLAFs were polymorphic, and 5,308 of the polymorphic markers met the requirements for use in constructing a genetic map. The final map included 5,308 markers on 20 linkage groups and was 2,655.68 cM in length, with an average distance of 0.5 cM between adjacent markers. To our knowledge, this map has the shortest average distance of adjacent markers for soybean. We report here a high-density genetic map for soybean. The map was constructed using a recombinant inbred line population and the SLAF-seq approach, which allowed the efficient development of a large number of polymorphic markers in a short time. Results of this study will not only provide a platform for gene/quantitative trait loci fine mapping, but will also serve as a reference for molecular breeding of soybean.

Increasing the yield of soybean (Glycine max L. Merrill) is a main aim of soybean breeding. The 100-seed weight is a critical factor for soybean yield. To facilitate genetic analysis of quantitative traits and to improve the accuracy of marker-assisted breeding in soybean, a valuable mapping population consisting of 194 chromosome segment substitution lines (CSSLs) was developed. In these lines, different chromosomal segments of the Chinese cultivar Suinong 14 were substituted into the genetic background of wild soybean (Glycine soja Sieb. & Zucc.) ZYD00006. Based on these CSSLs, a genetic map covering the full genome was generated using 121 simple sequence repeat (SSR) markers. In the quantitative trait loci (QTL) analysis, twelve main effect QTLs (qSW-B1-1/2/3, qSW-D1b-1/2, qSW-D2-1/2, qSW-G-1/2/3, qSW-M-2 and qSW-N-2) underlying 100-seed weight were identified in 2011 and 2012. The epistatic effects of pairwise interactions between markers were analyzed in 2011 and 2012. The results clearly demonstrated that these CSSLs could be used to identify QTLs, and that an epistatic analysis was able to detect several sites with important epistatic effects on 100-seed weight. Thus, we identified loci that will be valuable for improving soybean 100-seed weight. These results provide a valuable foundation for identifying the precise location of genes of interest, and for designing cloning and marker-assisted selection breeding strategies targeting the 100-seed weight of soybean.

Root nodules are major sources of nitrogen for soybean (Glycine max (L.) Merr.) growth, development, production, and seed quality. Symbiotic nitrogen fixation is time-limited, as the root nodule senesces during the reproductive stage of plant development, specifically during seed development. Nodule senescence is characterized by the induction of senescence-related genes, such as papain-like cysteine proteases (CYPs), which ultimately leads to the degradation of both bacteroids and plant cells. However, how nodule senescence–related genes are activated in soybean is unknown. Here, we identified two paralogous NAC transcription factors, GmNAC039 and GmNAC018, as master regulators of nodule senescence. Overexpression of either gene induced soybean nodule senescence with increased cell death as detected using a TUNEL assay, whereas their knockout delayed senescence and increased nitrogenase activity. Transcriptome analysis and nCUT&Tag-qPCR assays revealed that GmNAC039 directly binds to the core motif CAC(A)A and activates the expression of four GmCYP genes (GmCYP35, GmCYP37, GmCYP39 and GmCYP45). Similar to GmNAC039 and GmNAC018, overexpression or knockout of GmCYP genes in nodules resulted in precocious or delayed senescence, respectively. These data provide essential insights into the regulatory mechanisms of nodule senescence, in which GmNAC039 and GmNAC018 directly activate the expression of GmCYP genes to promote nodule senescence.

Transcriptional divergence of duplicated genes after whole genome duplication (WGD) has been described in many plant lineages and is often associated with subgenome dominance, a genome-wide mechanism. However, it is unknown what underlies the transcriptional divergence of duplicated genes in polyploid species that lack subgenome dominance. Soybean is a paleotetraploid with a WGD that occurred 5 to 13 Mya. Approximately 50% of the duplicated genes retained from this WGD exhibit transcriptional divergence. We developed accessible chromatin region (ACR) datasets from leaf, flower, and seed tissues using MNase-hypersensitivity sequencing. We validated enhancer function of several ACRs associated with known genes using CRISPR/Cas9-mediated genome editing. The ACR datasets were used to examine and correlate the transcriptional patterns of 17,111 pairs of duplicated genes in different tissues. We demonstrate that ACR dynamics are correlated with divergence of both expression level and tissue specificity of individual gene pairs. Gain or loss of flanking ACRs and mutation of cis -regulatory elements (CREs) within the ACRs can change the balance of the expression level and/or tissue specificity of the duplicated genes. Analysis of DNA sequences associated with ACRs revealed that the extensive sequence rearrangement after the WGD reshaped the CRE landscape, which appears to play a key role in the transcriptional divergence of duplicated genes in soybean. This may represent a general mechanism for transcriptional divergence of duplicated genes in polyploids that lack subgenome dominance.