Linkage Analysis For Sulfur-containing Amino Acid Related Traits And GmMS Gene Expression And Functional Study Of GmSET1 In Soybean

Soybean[Glycine Max(L.)Merr.]has a high protein content of about 40%.It is one of the most important sources of plant protein in the human diet and livestock breeding,but its amino acid content is not balanced,such as the content of sulfur-containing amino acids(methionine and cysteine),which limits the nutritional value of soybeans.Therefore,increasing the content of sulfur-containing amino acids in soybean seeds is the key to the improvement of soybean protein quality.The content of sulfur-containing amino acids in soybeans is mainly determined by genetic factors and is affected by environmental factors.It is a complex quantitative trait,which is regulated by multiple genes.Therefore,for the improvement of soybean nutritional quality,molecular marker-assisted selection(MAS)is more time-saving than traditional breeding methods.In this study,linkage analysis was performed for multiple traits about sulfur-containing amino acids and GmMS gene expression using two RIL populations of Kefeng and Bogo,which contained 184 and 146 lines,respectively.We identified QTLs and eQTLs associated with sulfur-containing amino acids,identified key QTL intervals,and screened for candidate genes of interest,the research results can provide new ideas for the cultivation of new high-sulfur amino acid varieties of soybean.The main research contents include:1.We determined Met/PC,Cys/PC,Met+Cys/PC(SAA/PC)and protein content(PC)of two RIL pop ulations,which constructed by "Ke fe ng No.1" and "Nannong 1 138-2",and"Nannong 94-156" and "Bogao",respectively.For the Kefeng RIL populations,using the value in 2017 and combing with the values in 2013 and 2014(Ma Yujie,2017)and their mean values to perform QTL analysis;for the Bogao RIL populations,using the value in 2017 and 2018 and their mean values to perform QTL analysis.Research shows:The four phenotypic values of each population exhibited a roughly consistent with normal distribution and an extensive variation.The correlation analysis showed that Cys/PC of each group was positively correlated with Met/PC and SAA/PC,and negatively correlated with PC.QTL analysis found that a total of 16 related QTLs were identified in the Kefeng RIL population,which including 3 major genomic segments distributed on chromosomes 8 and 18 respectively;a total of 18 related QTLs were identified in the Bogao RIL populations,which including 4 major genomic segments are distributed on chromosomes 8,13 and 16,respectively,which are the main genomic segments.2.We measured the expression levels of GmMS(methionine synthase gene)in 45 days of seeds after flowering and identified the eQTLs associated with it for Kefeng and Bogao RIL populations.The study found that the expression levels of GmMS showed extensive variation in Kefeng and Bogao RIL populations and the distribution was approximately normal.There are 8 eQTLs were identified in the Kefeng RIL populations,mainly distributed on chromosomes 7,9,12,13 and 18;a total of 17 eQTLs were identified in the Bogao RIL populations,mainly distributed on Chromosomes 2,3,5,6,7,9,10,13,15,16 and 19.However,the eQTLs associated with gene expression levels in the two populations were found to be located on different chromosomes or distantly from the genes regulated by the located eQTL,and no eQTLs were found that were physically close to the genome of the GmMS.Therefore,it is considered that the trans-eQTL is identified.It is hypothesized that the expression level of GmMS gene may only be regulated by trans-eQTL,or that the cis-eQTL that regulates its expression may not be identified in this study due to environmental factors.3.According to previous research results,a candidate gene(Glymal0g07420)was mapped to the chromosome 10 segment,and its homologous gene(AT2G36630)encodes sulfite export protein in Arabidopsis thaliana(Ma Yujie 2017),so we named it GmSET1.The full-length cDNA sequence of GmSET1 was cloned,expression analysis and transgene identification were carried out.Tissue expression analysis showed that GmSET1 was expressed in all tissues of soybean,and was higher in roots and young pods.It was overexpressed in soybean hairy roots and found that the content of Met and Cys in soybean hairy roots overexpressing GmSET1 was significantly increased,indicating that it is essential for the synthesis of soybean sulfur-containing amino acids.