Pan-Genome of Wild and Cultivated Soybeans.

Soybean is one of the most important vegetable oil and protein feed crops. To capture the entire genomic diversity, it is needed to construct a complete high-quality pan-genome from diverse soybean accessions. In this study, we performed individual de novo genome assemblies for 26 representative soybeans that were selected from 2,898 deeply sequenced accessions. Using these assembled genomes together with three previously reported genomes, we constructed a graph-based genome and performed pan-genome analysis, which identified numerous genetic variations that cannot be detected by direct mapping of short sequence reads onto a single reference genome. The structural variations from the 2,898 accessions that were genotyped based on the graph-based genome and the RNA sequencing (RNA-seq) data from the representative 26 accessions helped to link genetic variations to candidate genes that are responsible for important traits. This pan-genome resource will promote evolutionary and functional genomics studies in soybean.

Mutation of YL Results in a Yellow Leaf with Chloroplast RNA Editing Defect in Soybean.

RNA editing plays a key role in organelle gene expression. Little is known about how RNA editing factors influence soybean plant development. Here, we report the isolation and characterization of a soybean yl (yellow leaf) mutant. The yl plants showed decreased chlorophyll accumulation, lower PS II activity, an impaired net photosynthesis rate, and an altered chloroplast ultrastructure. Fine mapping of YL uncovered a point mutation in Glyma.20G187000, which encodes a chloroplast-localized protein homologous to Arabidopsis thaliana (Arabidopsis) ORRM1. YL is mainly expressed in trifoliate leaves, and its deficiency affects the editing of multiple chloroplast RNA sites, leading to inferior photosynthesis in soybean. Taken together, these results demonstrate the importance of the soybean YL protein in chloroplast RNA editing and photosynthesis.

The RING finger E3 ligase STRF1 is involved in membrane trafficking and modulates salt-stress response in Arabidopsis thaliana.

Salt stress is a detrimental factor for plant growth and development. The response to salt stress has been shown to involve components in the intracellular trafficking system, as well as components of the ubiquitin-proteasome system (UPS). In this article, we have identified in Arabidopsis thaliana a little reported ubiquitin ligase involved in salt-stress response, which we named STRF1 (Salt Tolerance RING Finger 1). STRF1 is a member of RING-H2 finger proteins and we demonstrate that it has ubiquitin ligase activity in vitro. We also show that STRF1 localizes mainly at the plasma membrane and at the intracellular endosomes. strf1-1 loss-of-function mutant seedlings exhibit accelerated endocytosis in roots, and have altered expression of several genes involved in the membrane trafficking system. Moreover, protein trafficking inhibitor, brefeldin A (BFA), treatment has increased BFA bodies in strf1-1 mutant. This mutant also showed increased tolerance to salt, ionic and osmotic stresses, reduced accumulation of reactive oxygen species during salt stress, and increased expression of AtRbohD, which encodes a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase involved in H2 O2 production. We conclude that STRF1 is a membrane trafficking-related ubiquitin ligase, which helps the plant to respond to salt stress by monitoring intracellular membrane trafficking and reactive oxygen species (ROS) production.

DNA methylation footprints during soybean domestication and improvement.

BACKGROUND: In addition to genetic variation, epigenetic variation plays an important role in determining various biological processes. The importance of natural genetic variation to crop domestication and improvement has been widely investigated. However, the contribution of epigenetic variation in crop domestication at population level has rarely been explored. RESULTS: To understand the impact of epigenetics on crop domestication, we investigate the variation of DNA methylation during soybean domestication and improvement by whole-genome bisulfite sequencing of 45 soybean accessions, including wild soybeans, landraces, and cultivars. Through methylomic analysis, we identify 5412 differentially methylated regions (DMRs). These DMRs exhibit characters distinct from those of genetically selected regions. In particular, they have significantly higher genetic diversity. Association analyses suggest only 22.54% of DMRs can be explained by local genetic variations. Intriguingly, genes in the DMRs that are not associated with any genetic variation are enriched in carbohydrate metabolism pathways. CONCLUSIONS: This study provides a valuable map of DNA methylation across diverse accessions and dissects the relationship between DNA methylation variation and genetic variation during soybean domestication, thus expanding our understanding of soybean domestication and improvement.

Assessing the fermentation quality and microbial community of the mixed silage of forage soybean with crop corn or sorghum.

The silage quality of forage soybean (FS) rich in protein with crop corn (CN) or sorghum (SG) rich in water soluble carbohydrate was investigated, and microbial community after ensiling was analyzed. Results showed that pH in mixed silages dropped to 3.5-3.8 lower than 100%FS silage (4.5). Microbial analysis indicated that mixed ensiling could influence the microbial community. Although Lactobacillus and Weissella were the dominant genera in all silage samples, Lactobacillus abundance in mixed silages (33-76%) was higher compared with 100%FS silage (27%). In conclusion, FS ensiled with CN or SG could be an alternative approach to improve FS silage quality.

Parallel selection on a dormancy gene during domestication of crops from multiple families.

Domesticated species often exhibit convergent phenotypic evolution, termed the domestication syndrome, of which loss of seed dormancy is a component. To date, dormancy genes that contribute to parallel domestication across different families have not been reported. Here, we cloned the classical stay-green G gene from soybean and found that it controls seed dormancy and showed evidence of selection during soybean domestication. Moreover, orthologs in rice and tomato also showed evidence of selection during domestication. Analysis of transgenic plants confirmed that orthologs of G had conserved functions in controlling seed dormancy in soybean, rice, and Arabidopsis. Functional investigation demonstrated that G affected seed dormancy through interactions with NCED3 and PSY and in turn modulated abscisic acid synthesis. Therefore, we identified a gene responsible for seed dormancy that has been subject to parallel selection in multiple crop families. This may help facilitate the domestication of new crops.

Effects of lactic acid bacteria and molasses additives on the microbial community and fermentation quality of soybean silage.

The objective was to study effects of lactic acid bacteria (L) and molasses (M) on the microbial community and fermentation quality of soybean silage. Soybean was ensiled with no additive control (C), 0.5% molasses (0.5%M), 0.5%M+L (0.5%ML), 2%M, 2%M+L (2%ML) for 7, 14, 30 and 60days. The M-treated silages could increase the content of lactic acid and decrease butyric acid than control. Besides, higher crude protein was also observed in M-treated silages. With prolonged ensiling time, there was a reduction of the ratio of lactic acid/acetic acid in the 2%M-treated and 2%ML-treated silages. The combined addition of L and 2%M could enhance the account of desirable Lactobacillus and inhibit the growth of undesirable microorganism such as Clostridia and Enterobacter. In summary, the silage quality of soybean was improved with the addition of L and M.

Genome-wide association studies dissect the genetic networks underlying agronomical traits in soybean.

BACKGROUND: Soybean (Glycine max [L.] Merr.) is one of the most important oil and protein crops. Ever-increasing soybean consumption necessitates the improvement of varieties for more efficient production. However, both correlations among different traits and genetic interactions among genes that affect a single trait pose a challenge to soybean breeding. RESULTS: To understand the genetic networks underlying phenotypic correlations, we collected 809 soybean accessions worldwide and phenotyped them for two years at three locations for 84 agronomic traits. Genome-wide association studies identified 245 significant genetic loci, among which 95 genetically interacted with other loci. We determined that 14 oil synthesis-related genes are responsible for fatty acid accumulation in soybean and function in line with an additive model. Network analyses demonstrated that 51 traits could be linked through the linkage disequilibrium of 115 associated loci and these links reflect phenotypic correlations. We revealed that 23 loci, including the known Dt1, E2, E1, Ln, Dt2, Fan, and Fap loci, as well as 16 undefined associated loci, have pleiotropic effects on different traits. CONCLUSIONS: This study provides insights into the genetic correlation among complex traits and will facilitate future soybean functional studies and breeding through molecular design.

[Cloning of GmHSFA1 gene and its overexpression leading to enhancement of heat tolerance in transgenic soybean].

Heat shock transcription factors (HSFs) are important in regulating heat stress response by mediating expression of heat shock protein (HSP) genes in various plant species. In the present study, a novel GmHSFA1 with an ORF of 1,533 bp (full-length cDNA sequence of 1,781 bp) was cloned from soybean genome via comparative genomic approach and RACE (rapid amplification of cDNA ends). This gene encodes 510 amino acids consisting of a protein of 56.2 kDa (GenBank accession number: AY458843). Similar to other HSFs, GmHSFA1 has the basic modular structure including DBD, OD, NLS, and CTAD. BLAST analysis revealed the identity of 52.46% between amino acid sequences between GmHSFA1 and LpHSFA1 that has the highest similarity to GmHSFA1 in all HSFA1s in various plant species. The results from RT-PCR, Northern blotting, and transformation showed: 1) GmHsfA1 exhibited the constitutive expression patterns in different tissues of soybean; 2) The expression level of GmHsfA1 in transgenic plants was notably higher than that in non-transgenic plants; 3) Overexpression of GmHsfA1 activated transcription of GmHSP22 in transgenic plants under normal conditions and enhanced obviously expressions of GmHSP23 and GmHSP70 in transgenic plants under heat stress conditions; 4) Heat tolerant temperature (as high as 52 degrees C) of transgenic plants was remarkably higher than that of non-transgenic plants. These results preliminarily proved that the overexpression of GmHsfA1 possibly led to the notable enhancement of heat-tolerant level of transgenic plants by mediating the activation of transcription or improvement of expression of some GmHSPs in the GmHsfA1's downstream in transgenic plants, suggesting GmHSFA1 is a novel and functional heat shock transcription factor of soybean.

Resequencing 302 wild and cultivated accessions identifies genes related to domestication and improvement in soybean.

Understanding soybean (Glycine max) domestication and improvement at a genetic level is important to inform future efforts to further improve a crop that provides the world's main source of oilseed. We detect 230 selective sweeps and 162 selected copy number variants by analysis of 302 resequenced wild, landrace and improved soybean accessions at >11× depth. A genome-wide association study using these new sequences reveals associations between 10 selected regions and 9 domestication or improvement traits, and identifies 13 previously uncharacterized loci for agronomic traits including oil content, plant height and pubescence form. Combined with previous quantitative trait loci (QTL) information, we find that, of the 230 selected regions, 96 correlate with reported oil QTLs and 21 contain fatty acid biosynthesis genes. Moreover, we observe that some traits and loci are associated with geographical regions, which shows that soybean populations are structured geographically. This study provides resources for genomics-enabled improvements in soybean breeding.

Evolutionary analysis of three gibberellin oxidase genes in rice, Arabidopsis, and soybean.

GAs are plant hormones that play fundamental roles in plant growth and development. GA2ox, GA3ox, and GA20ox are three key enzymes in GA biosynthesis. These enzymes belong to the 2OG-Fe (II) oxygenase superfamily and are independently encoded by different gene families. To date, genome-wide comparative analyses of GA oxidases in plant species have not been thoroughly carried out. In the present work, 61 GA oxidase family genes from rice (Oryza sativa), Arabidopsis, and soybean (Glycine max) were identified and a full study of these genes including phylogenetic tree construction, gene structure, gene family expansion and analysis of functional motifs was performed. Based on phylogeny, most of the GA oxidases were divided into four subgroups that reflected functional classifications. Intron/intron average length of GA oxidase genes in rice analysis revealed that GA oxidase genes in rice experienced substantial evolutionary divergence. Segmental duplication events were mainly found in soybean genome. However, in rice and Arabidopsis, no single expansion pattern exhibited dominance, indicating that GA oxidase genes from these species might have been subjected to a more complex evolutionary mechanism. In addition, special functional motifs were discovered in GA20ox, GA3ox, and GA2ox, which suggested that different functional motifs are associated with differences in protein function. Taken together our results suggest that GA oxidase family genes have undergone divergent evolutionary routes, especially at the monocot-dicot split, with dynamic evolution occurring in Arabidopsis thaliana and soybean.

Identification and characterization of a novel heat shock transcription factor gene, GmHsfA1, in soybeans (Glycine max).

Plants have a large family of HSFs with different roles in the heat shock response that mediate the expression of HSP regulated genes. The HSF encoding genes are easily identified by their highly conserved modular structure and motifs. In the present study, a putative GmHsfA1 was identified and characterized from the soybean expressed sequence tag (EST) database by sequence comparison with the functionally well-characterized LpHsfA1 and rapid amplification of cDNA ends (RACE). Multiple alignment showed that the amino acid sequence of GmHSFA1, matching best with LpHSFA1 (52.2% similarity), was obviously different from that of each of several HSFA1s from other plant species. The GmHsfA1 has a constitutive expression profile in the different tissues examined. The overexpression of GmHsfA1 in transgenic soybean plants led to the activation of GmHsp70 under normal temperature and the overexpression of GmHsp70 under high temperature. Furthermore, transgenic soybean plants with GmHsfA1 overexpression showed obvious enhancement of thermotolerance under heat stress in comparison with non-transgenic plants. The experimental results suggested that GmHSFA1 is a novel and functional heat-shock transcription factor.