Wheat Improvement With Two Quality-associated HMW-GS Genes (H1Dx5and H1Bx13) By Agrobacterium-mediated Transformation

Common wheat(Triticnm aestivum L.)is one of the most important crops in the world. High-Molecular-Weight glutenin subunits(HMW-GS) are important determents High molecular weight (HMW) glutenin subunits are important determinants of the baking quality of bread wheat, and the presence of certain HMW glutenin subunits have different association with wheat bread making quality.Up to now, more and more HMW-GS genes with good bread making quality have been cloned due to the improvement of gene engineering. In many of the recent experiments, researchers transform these genes for high quality into those wheat varieties that have high output but low quality, which could improve the quality of the accepter wheat.Asymmetric somatic hybridization between protoplast of common wheat Jinan177and UV-irradiated protoplast of Agropyron elongatum generated fertile introgression hybrid lines, from which many novel HMW-GS genes with good quality were cloned. In this research we chose two genes called H1Dx5and H1Bx13They were finally constructed into the reformed plant transformation vector p3301-1forming three new vectors named p3301-1-HlBxl3,p3301-1-HlDx5,P3301-1-HlDx5-HlBx13.Each gene was linked with specificity promoter1Bx17.Several wheat varieties of Henan and Shandong,including Shanrong No.3,Jimai22,Zhengmai366,Zhoumai18,Zhoumai22and07YT2122et al, were transformed by dipping dipping shoot tjp with Agrobatenum strain of EHA105harboring these vectors constructed. Gus gene was used for selecting positive transgenic generations. We finally get plenties of TO and T1generation. SDS-PAGE analysis indicated that among171TO and100T1generation seedings there is no expected protein expressed. However, the certain HMW-GS in some of these transgenic seedings are lost with the possible reason that the foreign genes influence the expression of the endogenous genes.In addition, we also cloned the promoters of HMW-GS genes1Ax1,1Bx7,1Bx14,1Byl5,1Dx5and1Dy10from several wheat variaties. All of these promoters’sequences cloned are more than2.0kb long. These sequences contains E-box,N-box,G-box, HMW-GS specific38bp enhancer and TATA-box, which were promoter role regulatory elements of the typical HMW-GS gene. We constructed these promoters into two expression vectors pSTART and pGA3383respectively, and some of the new vectors were transformed into Arabidopsis thaliana (Col-0) and Brachypodium distachyon (Bd21).In the further research, we will select the stable express transplants, and all the transgenic lines will be used to verify the activity and efficiency of these HMW-GS promoters cloned in this experiment. The present results provide many effective information for HMW-GS gene transformation and wheat quality improvement.