Summary: Isoflavonoids, which include a variety of secondary metabolites, are derived from the phenylpropanoid pathway and are distributed predominantly in leguminous plants. These compounds play a critical role in plant–environment interactions and are beneficial to human health. Isoflavone synthase (IFS) is a key enzyme in isoflavonoid synthesis and shares a common substrate with flavanone‐3‐hydroxylase (F3H) and flavone synthase II (FNS II). In this study, CRISPR/Cas9‐mediated multiplex gene‐editing technology was employed to simultaneously target GmF3H1 , GmF3H2 and GmFNSII‐1 in soya bean hairy roots and plants. Various mutation types and frequencies were observed in hairy roots. Higher mutation efficiencies were found in the T ₀ transgenic plants, with a triple gene mutation efficiency of 44.44%, and these results of targeted mutagenesis were stably inherited in the progeny. Metabolomic analysis of T ₀ triple‐mutants leaves revealed significant improvement in isoflavone content. Compared with the wild type, the T ₃ generation homozygous triple mutants had approximately twice the leaf isoflavone content, and the soya bean mosaic virus (SMV) coat protein content was significantly reduced by one‐third after infection with strain SC7, suggesting that increased isoflavone content enhanced the leaf resistance to SMV. The isoflavone content in the seeds of T ₂ triple mutants was also significantly increased. This study provides not only materials for the improvement of soya bean isoflavone content and resistance to SMV but also a simple system to generate multiplex mutations in soya bean, which may be beneficial for further breeding and metabolic engineering.

Previous studies suggest that the metabolic pathway structure influences the selection and evolution rates of involved genes. However, most of these studies have exclusively considered a single gene copy encoding each enzyme in the metabolic pathway. Considering multiple-copy encoding enzymes could provide direct evidence of gene evolution and duplication patterns in metabolic pathways. We conducted a detailed analysis of the phylogeny, synteny, evolutionary rate and selection pressure of the genes in the isoflavonoid metabolic pathway of soybeans.

Background: Phosphorus (P) is an essential element in maintaining high biomass and yield in crops. Soybean [Glycine max (L.) Merr.] requires a large amount of P during growth and development. Improvement of P efficiency and identification of P efficiency genes are important strategies for increasing soybean yield. Results: Genome-wide association analysis (GWAS) with NJAU 355 K SoySNP array was performed to identify single nucleotide polymorphisms (SNPs) significantly associated with three shoot P efficiency-related traits of a natural population of 211 cultivated soybeans and relative values of these traits under normal P (+P) condition and P deficiency (−P) condition. A total of 155 SNPs were identified significantly associated with P efficiency-related traits. SNPs that were significantly associated with shoot dry weight formed a SNP cluster on chromosome 11, while SNPs that were significantly associated with shoot P concentration formed a SNP cluster on chromosome 10. Thirteen haplotypes were identified based on 12 SNPs, and Hap9 was considered as the optimal haplotype. Four SNPs (AX-93636685, AX-93636692, AX-93932863, and AX-93932874) located on chromosome 10 were identified to be significantly associated with shoot P concentration under +P condition in two hydroponic experiments. Among these four SNPs, two of them (AX-93636685 and AX-93932874) were also significantly associated with the relative values of shoot P concentration under two P conditions. One SNP AX-93932874 was detected within 5′-untranslated region of Glyma.10 g018800, which contained SPX and RING domains and was named as GmSPX-RING1. Furthermore, the function research of GmSPX-RING1 was carried out in soybean hairy root transformation. Compared with their respective controls, P concentration in GmSPX-RING1 overexpressing transgenic hairy roots was significantly reduced by 32.75% under +P condition; In contrast, P concentration in RNA interference of GmSPX-RING1 transgenic hairy roots was increased by 38.90 and 14.51% under +P and -P conditions, respectively. Conclusions: This study shows that the candidate gene GmSPX-RING1 affects soybean phosphorus efficiency by negatively regulating soybean phosphorus concentration in soybean hairy roots. The SNPs and candidate genes identified should be potential for improvement of P efficiency in future soybean breeding programs.