Identification Of High Folate Foxtail Millet Germplasm And Characterization,Expression And Genetic Transformation Of SiFPGS Gene

Millet,after the foxtail millet seeds were husked,is known as the "the best grain." Due to its seeds contained many nutritional Components,many scholars were attracted in studying its nutritional value.Folate is one of the vitamins B,participated in transferring one-carbon units in cells and regulating embryonic neural cell development.However,a few researches on folic acid of foxtail millet were reported and published.Based on chemical extraction methods,the folic acid content of 45 foxtail millet germplasm resources were detected by the high performance liquid chromatography(HPLC)methods.were used and analyzed o differences in folic acid content among various foxtail millet varieties.From the molecular level,two FPGS gene named FPGS1 and FPGS2,in foxtail millet were cloned and carried out quantitative gene expression analysis.However,,overexpressed vector of FPGS 1 were constructedand transferred into Arabidopsis thaliana,as a key enzyme in the folic acid synthesis pathway.The aim is to reveal the molecular mechanism of folic acid synthesis and lay a theoretical foundation for breeding of high folic acid foxtail millet varieties in the future.The major conclusions are stated as follows:(1)Using high performance liquid chromatography(HPLC)to determine the folate content of 45 foxtail millet germplasm resources:the folic acid content of 45 foxtail millet varies were ranged from 0.23 ?g.g-1 to 2.68?g·g-1,and the average content of folic acid was 1.52?g·g-1,and the coefficient of variation is 38.8%.Among them,the varieties with high folate content were 'Yugu 1' 'Jinfen 52','Jingu 35','Jingu 29','Changsheng 08','Jingu 21',etc,and the varieties with low folate content include 'Pingdingdagu',etc.(2)SiFPGS genes was cloned from' Changsheng 08' and ' Pingdingdagu',finally,two SiFPGS genes were obtained.Evolutionary analysis of homologous genes from other species shows that they belong to two homologous genes,named SiFPGS-1 and SiFPGS-2,respectively.SiFPGS-1,the cDNA sequence is 1248 bp and encodes 415 amino acids.SiFPGS-2,the cDNA sequence is 1740bp,encodes 579 amino acids.The conserved domain analysis of protein sequence showed that all of them had the function of polyglutamic acid synthase.(3)Analysis of the folic acid content in the tissues of 'Changsheng 08' and'Pingdingdagu' the whole growth period:In leaves,stems,flower tissue,the content of folic acid in leaves is the highest.With the grain from filling to full maturity,the folic acid content showed a trend of increasing first and then decreasing.In addition,the correlation analysis of SiFPGS gene expression pattern and folic acid content showed that the correlation was not significant.(4)Constructed SiFPGS-1 overexpression vector and genetically transformed Arabidopsis thaliana.93 transgenic lines were screened from 2189 T1 generation lines,and the positive rate was 4.2%.