Soybean Flowering Time Regulation And Protein Quality Improvement Based On The Constitutive Expression Of GmFT2a And AtD-CGS

Soybean(Glycine max(L)Merr.)is one of the most widely-grown field crops in the world.Nowadays,the major transgenic trait is herbicide resistantance in genetic modified(GM)soybean.However,coming demands from soybean molecular breeding were growing fast.To solve the problems that the convention breeding hardly coped with,we focused on two major issues druing the production of the soybean,which including light/temperature sensitivity caused adaptation limitation and the scarcity of sulfur-contain amino acid(such as,methionine and cysteine)in soybean cultivars,and attempted to designed molecular breeding strategies to solved this problems.Supporting by the stable transformation platform of the soybean,we modified the upper two traits by expression key genes GmFT2 a and AtD-CGS in the flowering integrate pathway and in methionine synthesis pathway in order to control the flowering time and improve the soybean protein quality resepctively.The transgenic progenies of above these transgenic soybean lines provided us superior materials to put further research on related mechanisms.Aiming to control the soybean flowering time,we obtained early flowering transgenic soybeans by overexpressing Gm FT2 a in a late flowering variety Zigongdongdou.The trasncriptional analysis showed that early flowering phenotype directly caused by the expression of Gm FT2 a,but not the other floral genes in transgenic progenies.Under different photoperiod conditions,we found that the early flowering phenotypes of the transgenic progenies demostrated a dose-wise effect.Thus,we subsequently detected the expression level in transgenic soybeans with diverged flowering times.The result showed that the expression level of Gm FT2 a correlated to the early flowering days among the transgenic progenies.Moreover,the early flowering phenotype in transgenic progenies also changed the structure of growth period and led to lower height and yield of the plant.Unstable flowering phenotype was found among the generations and the offspring individuals.The copy number variation(CNV)in transgenic soybeans showed that multiple copies in plant may lead to expression inhibition and limit the early flowering.A 129 bp CpG island was found in promoter region and the methylated percentage in this region of promoter highly explains the correlation with the diversed flowering time in transgenic progenies.According to our result from the transcriptome analysis and qPCR detection,it indicated that both the transgenic soybeans and the wild type could affect by the expression inhibition specifically on GmFT2 a and led to a lower level in the former development period.In addition,through the transcriptome analysis,we identified three differential expression genes including AGL-8,PRR,and SPA1,which highly related to flowering regulation,between the transgenic and wild type soybeans.To increase the methionine content in soybean,we firstly analyzed the GmCGS gene and the protein structure with several other reported plants.It suggested that a functional region in GmCGS may response to the feed-back inhibition of the excess methionine.According to our result,the GmCGS expression was indeed suppressed when the leaves was treated by methionine spraying.Hence,it led us to express a feed-back insensitive gene AtD-CGS to enhance the synthesis of the methionine in soybean.The transgenic soybeans displayed stable expression in soybean leaf and seed both in transcriptional and translational level.However,in some transgenic lines the dwarf individuals were observed.We further detected the methionine content in transgenic soybean.The results demonstrated that the soluble methionine content was elevated in leaves,whereas total content of methionine was not altered in leaves.Total methionine in transgenic soybeans was significantly elevated in seeds,whereas soluble methionine showed different alternations between the two cultivars Zigongdongdou and Jilinxiaoli No.1.It also showed that the increasing of methionine content led to a higher accumulation in seed protein and a decreasing content in lipid.Comparing to our former result,it suggested that the soluble methionine content may act as the bottle neck that limited the total methionine and protein accumulation in soybean seeds.In general,it is a practicable way to regulate the flowering time and improve the soybean quality via designed molecular breeding strategies in this study.Moreover,it could also provide valuable materials for the researches on related mechanisms and lay a foundation for molecular breeding of soybean.