SoyFGB v2.0: a unique access to variations of Chinese Soybean Gene Bank (CNSGB) germplasm

In Chinese National Soybean GeneBank (CNSGB), we have collected more than 30,000 soybean accessions. However, data sharing for soybean remains an especially sensitive question, and how to share the genome variations within rule frame has been bothering the soybean germplasm workers for a long time. Here we release a big data source named Soybean Functional Genomics & Breeding database (SoyFGB v2.0) (https://sfgb.rmbreeding.cn/), which embed a core collection of 2,214 soybean resequencing genome (2K-SG) from the CNSGB germplasm. This source presents a unique example which may help elucidating the following three major functions for multiple genome data mining with general interests for plant researchers. 1) On-line analysis tools are provided by the Analysis module for haplotype mining in high-throughput genotyped germplasms with different methods. 2) Variations for 2K-SG are provided in SoyFGB v2.0 by Browse module which contains two functions of SNP and InDel. Together with the Gene (SNP & InDel) function embedded in Search module, the genotypic information of 2K-SG for targeting gene / region is accessible. 3) Scaled phenotype data of 42 traits, including 9 quality and 33 quantitative traits are provided by SoyFGB v2.0. With the scaled-phenotype data search and seed request tools under a control list, the germplasm information could be shared without direct downloading the unpublished phenotypic data or information of sensitive germplasms. In a word, the mode of data mining and sharing underlies SoyFGB v2.0 may inspire more ideas for works on genome resources of not only soybean but also the other plants.

Expression of an Antiviral Gene GmRUN1 from Soybean Is Regulated via Intron-Mediated Enhancement (IME)

Most of R (resistance) genes encode the protein containing NBS-LRR (nucleotide binding site and leucine-rich repeat) domains. Here, N. benthamiana plants were used for transient expression assays at 3–4 weeks of age. We identified a TNL (TIR-NBS-LRR) encoding gene GmRUN1 that was resistant to both soybean mosaic virus (SMV) and tobacco mosaic virus (TMV). Truncation analysis indicated the importance of all three canonical domains for GmRUN1-mediated antiviral activity. Promoter-GUS analysis showed that GmRUN1 expression is inducible by both salicylic acid (SA) and a transcription factor GmDREB3 via the cis-elements as-1 and ERE (ethylene response element), which are present in its promoter region. Interestingly, GmRUN1 gDNA (genomic DNA) shows higher viral resistance than its cDNA (complementary DNA), indicating the existence of intron-mediated enhancement (IME) for GmRUN1 regulation. We provided evidence that intron2 of GmRUN1 increased the mRNA level of native gene GmRUN1, a soybean antiviral gene SRC7 and also a reporter gene Luciferase, indicating the general transcriptional enhancement of intron2 in different genes. In summary, we identified an antiviral TNL type soybean gene GmRUN1, expression of which was regulated at different layers. The investigation of GmRUN1 gene regulatory network would help to explore the mechanism underlying soybean-SMV interactions.

Extraction of Damba Black Soybean Gene by Means of Magnetic Nanoparticles and Investigation of Aluminum Tolerance

The aluminum tolerance of Tamba black soybean (Glycine max cv. Tamba) is closely related to organic acid secretion mechanisms. The gene responsible for this tolerance in this variety (GmFER84) is extracted from lysates of soybean root tips using silylated Fe3O4 nanomagnetic beads. GmFER84 (Glycine max XP 003540203.1) is a stable protein. Tobacco genetically transformed with GmFER84 using an Agrobacterium-mediated transformation was tested for aluminum tolerance. Citrate synthase and citric acid secretion in the roots of transgenic tobacco prove to be significantly higher than those of wild tobacco, and the antioxidant properties of transgenic tobacco are also substantially increased. Research on GmFER84 may enable further agronomic development.

GmNMHC5, A Neoteric Positive Transcription Factor of Flowering and Maturity in Soybean

The soybean (Glycine max (L.) Merr.) is an important oil and food crop. Its growth and development is regulated by complex genetic networks, and there are still many genes with unknown functions in regulation pathways. In this study, GmNMHC5, a member of the MADS-box protein family, was found to promote flowering and maturity in the soybean. Gene expression profiling in transgenic plants confirmed that the 35S:GmNMHC5 T3 generation had early flowering and precocity. We used CRISPR-Cas9 to edit GmNMHC5 and found that late flowering and maturity occurred in Gmnmhc5 lines with stable inheritance. Remarkably, in the 35S:GmNMHC5 plants, the expression of flowering inhibitors GmFT1a and GmFT4 was inhibited. In addition, overexpression of GmNMHC5 in ft-10 (a late flowering Arabidopsis thaliana mutant lacking Flowering Locus T (FT) function) rescued the extremely late-flowering phenotype of the mutant A. thaliana. These results suggest that GmNMHC5 is a positive transcription factor of flowering and maturity in the soybean, which has a close relationship with FT homologs in the flowering regulation pathway. This discovery provides new ideas for the improvement of the flowering regulation network, and can also provide guidance for future breeding work.

Heterologous Expression of a Soybean Gene RR34 Conferred Improved Drought Resistance of Transgenic Arabidopsis

Two-component systems (TCSs) have been identified as participants in mediating plant response to water deficit. Nevertheless, insights of their contribution to plant drought responses and associated regulatory mechanisms remain limited. Herein, a soybean response regulator (RR) gene RR34, which is the potential drought-responsive downstream member of a TCS, was ectopically expressed in the model plant Arabidopsis for the analysis of its biological roles in drought stress response. Results from the survival test revealed outstanding recovery ratios of 52%–53% in the examined transgenic lines compared with 28% of the wild-type plants. Additionally, remarkedly lower water loss rates in detached leaves as well as enhanced antioxidant enzyme activities of catalase and superoxide dismutase were observed in the transgenic group. Further transcriptional analysis of a subset of drought-responsive genes demonstrated higher expression in GmRR34-transgenic plants upon exposure to drought, including abscisic acid (ABA)-related genes NCED3, OST1, ABI5, and RAB18. These ectopic expression lines also displayed hypersensitivity to ABA treatment at germination and post-germination stages. Collectively, these findings indicated the ABA-associated mode of action of GmRR34 in conferring better plant performance under the adverse drought conditions.

Design of Construct Carrying GmDREB6 to Enhance Soybean Gene Expression Related to Abiotic Stress Response

Glycine max (L.) Merrill is a crop that brings a lot of economic and nutritive values, however soybean is quite sensitive to stress. The applications of gene technology can improve resistance of soybean plants against external stress factors. The aim of this study was to conduct a transgenic vector containing GmDREB6 gene and determine the expression of gene encoding GmDREB6 in Nicotiana tabacum before transforming into soybean plants. The GmDREB6 artificial gene was synthesized containing nucleotide fragment encoding 230 amino acids, nucleotide fragment encoding cmyc antigen and nucleotide fragments with cut-off points of XbaI/SacI enzyme pair. The results indicated that the 35S-GmDREB6-cmyc construct was designed and transferred into tobacco plants. The GmDREB6 was incorporated in the genome and was expressed in transgenic tobacco plants at the transcriptional level. The transgenic vector pBI121_GmDREB6 well worked on the model tobacco plants. Therefore, it can be used for transferring into soybean plants to enhance soybean tolerance to abiotic stress.

Characteristics of mutant soybean lines at Satt100 and Satt319 loci linked with E7 gene

Aim. Аnalysis of genetic polymorphism of microsatellite loci Satt100 and Satt319 in 10 lines obtained by chemical mutagenesis, and 4 parental varieties – Oksana, Femida, Zolotysta, Podil’s’ka 416. Satt100 and Satt319 are flanking E7gene, which determines sensitivity of soybean to the length of the day in ripening phase. Methods. DNA were isolated from soybean seeds using the kit NeoPrep100 DNA. PCR were performed with microsatellites Satt100 and Satt319. The PCR products were fractionated in 7% polyacrylamide gels. Results. In the investigated lines were detected 5 alleles at the Satt100 locus and 3 alleles at the Satt319 locus. 42.9 % of the varieties and lines in this study were carriers of the «E» allele for Satt100. According to analysis of locus Satt319 64.2 % of varieties and lines were the carriers of «B» allele. Conclusions. Variety Oksana, mutant lines Oksana M №12, Femida M №29 – are carriers of the dominant allele E7. Variety Zolotysta and five mutant lines have recessive allele e7. In 75 % cases we have detected changes of alleles at microsatellite loci in mutant lines in comparison with parental forms. We assume that mutagenic reagents could affect at generative organs of soybean and lead totheir open flowering and crosspollinations. Keywords: Glycine max (L.), soybean, gene E7, microsatellite markers, photoperiodic sensitivity.

Whole-Genome Resequencing Identifies the Molecular Genetic Cause for the Absence of a Gy5 Glycinin Protein in Soybean PI 603408

During ongoing proteomic analysis of the soybean (Glycine max (L.) Merr) germplasm collection, PI 603408 was identified as a landrace whose seeds lack accumulation of one of the major seed storage glycinin protein subunits. Whole genomic resequencing was used to identify a two-base deletion affecting glycinin 5. The newly discovered deletion was confirmed to be causative through immunological, genetic, and proteomic analysis, and no significant differences in total seed protein content were found to be due to the glycinin 5 loss-of-function mutation per se. In addition to focused studies on this one specific glycinin subunit-encoding gene, a total of 1,858,185 nucleotide variants were identified, of which 39,344 were predicted to affect protein coding regions. In order to semiautomate analysis of a large number of soybean gene variants, a new SIFT 4G (Sorting Intolerant From Tolerated 4 Genomes) database was designed to predict the impact of nonsynonymous single nucleotide soybean gene variants, potentially enabling more rapid analysis of soybean resequencing data in the future.