The Primary Study On Expresssion Of LMW-GS Gene XYGluD3-LMWGS1 From Common Wheat In Pichia Pastoris

Wheat is one of the most important crops in the world, and the plant area of wheat is about 30% of all crops. The flour can be machininged all kinds of foodstuff and it is one of the sources of proteins for mankind.Therefore on the study of wheat storage protein is one of the focuses in cereal science.Both high- and low-molecular-weight glutenin subunits are the important components of wheat seed storage. Triticum aestivum low-molecular-weight glutenins subunits (LMW-GS) is a kind of important storage protein and account for 40% of wheat gluten protein content. These proteins play a significant role on flour-processing properties. LMW and High-molecular-weight glutenins subunits (HMW-GS) are considered to significantly affect dough-quality characteristics. They incorporate in glutenin macropolymer (GMP) through intermolecular disulphide bonds. It was reported that the content,component and size distribution significantly affect the quality characteristics of wheat flour.The composition and quanity of glutenins affect the size distribution of glutenin macropolymer (GMP). So, wheat flour processing properties can be improved by enhancing the quanity of GMP and altering GMP component through introducing genes related to good quality into wheat cultivar.This reasearch amplified XYGluD3-LMWGS1 gene by PCR,and constructed the yeast expressive vectors.The expression of XYGluD3-LMWGS1 in Pichia.pasteoris were studied.The main achievements were as follows:1.XYGluD3-LMWGS1 gene was amplified from the pGEM-XYGluD3-LMWGS1,the XYGluD3-LMWGS1 gene was cloned into PMD18-simple T.The recombinants were assayed by the digestion of restricted enzyme and sequenced.The results indicated that the XYGluD3-LMWGS1 gene was cloned into PMD18-simple T.2.The clone vector and pPIC3.5K were digested by the restricted enzyme,the ligated by T4 DNA ligase.The XYGluD3-LMWGS1 gene was cloned into pPIC3.5K by analysis.3. The expressive vector was transformed into Pichia.pasteoris(GS115) by electricity dash method.Transformants were obtained by PCR.The recombinants were obtained bt the assay of PCR.4.The strain express the XYGluD3-LMWGS1 in the culture medium using methyl alcoholas the carbolic rescource.Through the SDS-PAGE,expressed pretein were not found.Analysis of the sequence of gene and the condition of culture were found,the optimal condon just 58% in all condon,not enough optimal condon maybe influence the efficiency of translation bring down.The PH,temperature and other conditions maybe influence the expression of protein.