酵母双杂诱饵表达载体pGBKT7-ufgt的构建与表达  

Construction and Expression of Yeast Two-hybrid Bait Expression Vector pGBKT7-ufgt

作　　者：董燕梅[1] 张鹏飞[1] 张小军[1] 高燕[1] 温鹏飞[1] 

机构地区：[1]山西农业大学园艺学院,太谷030801

出　　处：《分子植物育种》2018年第4期1147-1153,共7页Molecular Plant Breeding

基　　金：国家自然科学基金项目(No.31372013);山西省科技重点研发(指南)项目(No.201603D211105-8);山西省科技成果转化项目(No.201604D132034)共同资助

摘　　要：构建葡萄赤霞珠(Vitis vinifera L.cv Cabernet Sauvignon)花色苷合成前体酶——类黄酮葡萄糖基转移酶(UDPG-flavonoid-3-O-glycosyltranferase,Ufgt)诱饵表达载体。通过RT-PCR扩增获得葡萄ufgt基因CDS序列,与p GBKT7酵母诱饵载体连接;经测序和双酶切鉴定后,转化酵母菌株AH109,分析Ufgt蛋白在酵母菌株中的表达情况。成功扩增出ufgt基因CDS全长,并成功构建了p GBKT7-ufgt,重组载体p GBKT7-ufgt成功的转化到AH109酵母菌株中,且无毒和自激活性,能够正确稳定的表达。p GBKT7-ufgt酵母诱饵蛋白表达载体被成功构建,为后续Ufgt蛋白互作筛选奠定基础。The bait expression plasmid of pGBKT7-ufgt was constructed. The ufgt fragment was amplified by PCR and cloned into the bait vector of pGBKT7. After identified by DNA sequencing and double enzyme, the combinant plasmid was transformed into AH109 yeast cells, ufgt expression was then analyzed. The ufgt fragment was successfully amplified and cloned into pGBKT7. The bait expression vector was transformed into AH109 yeast cells. Ufgt protein was expressed in the bait vector, without toxicity and self-activation. The bait plasmid of pGBKT7-ufgt was success fully constructed using yeast two-hybrid technique.

关 键 词：ufgt 诱饵蛋白 酵母双杂交 

分 类 号：Q78[生物学—分子生物学]