抗病小麦中与大麦黄矮病毒CP蛋白和RdRp复制酶相互作用蛋白的研究-作物遗传育种专业毕业论文.docx

AbstractWheat Abstract Wheat yellow dwarf caused by barley yellow dwarf virus(BYDV)is one of the most itnportant diseases of wheat worldwide．Genes resistant to BYDV originated from Thinopyrura intermedium have been introduced into the genuine of hexaploid whv．．钆and S01Ee BYDV resistant materials such as HW642 have been got in our research group．The BYDV resistant genes from HW642 Call inhibit the replication and movement of BYDV．In order to know the wheat protein interacting with the proteins of BYDV,yeast two hybrid system was used tO isolate the protein interacting with BYDV coat protein (CP)and RNA-dependent RNA polymerase(RdRp)from the eDNA library of HW642．The structure of the candidate gone was to be predicted,and its expression model would be analyzed．The result ofthe present research wag shown嬲the following． 1．Yeast two hybrid bait vectors of coat protein(CP)and RNA-dependent RNA polymerase (RdRp)of barley yellow dwarf virus(BYDV)were constructed and expressed in yeast．Yeast containing bait VeCtor pGBKTT-CP and pGBKT／-RdRp could grow On SD／-Trp culture medium， but couldn’t grow锄SD／-Ade／-Trp／X-Gai and SD／-Sis／-rrpm-C谢mediunl．ne result indicated the pGBKT7-CP and pGBKT7一RdRp had no toxicity and autonomous activation in yeast,and could be used as bait veCtor in yeast two hybrid system to analyze the proteins interacted with CP and RdRp ofBYDV． 2．A yeast two hybrid eDNA library of HW642 Was constructed(MatchmakerTM Library Construction＆ScreeningKits．Clotech)．n碹librarycomprisederatotalofsoo,000colonies．The library titer is 3x107 a删。recombinant rate is about 88％，and the inserted fragment ranges from0．2to 1．4kb． 3．To idantify the host proteins that interact with BYDV CP,yeast two-hybrid system was used to screen a wheat cDNA library．Using full—length BYDV CP as bait，a candidate eDNA clone was isolated,designated TaSTKl．TaSTKl encodes a protein shares high homogeneity with serine／threonine-protein kinase(STKl．The further e—gaiactosidase quantitative assay and veCt