Asp896～Val911缺失的阴离子交换蛋白1C末端表达质粒的构建

The construction of anion exchange 1 C-terminal expression plasmid with Asp896～Val911 deletion

目的 构建Asp896～Val911缺失的阴离子交换蛋白 1(AnionExchange1,AE1)C末端表达质粒。方法 利用PCR方法 ,以pFAST Bac AE1 C end为模板 ,扩增出AE1后 16位氨基酸Asp896～Val911缺失的C末端基因 ,将其克隆至pGADT7载体上。用醋酸锂法将构建好的pGADT7 AE1 c end truncation转染酵母菌AH10 9,观察其在选择性培养基上的表达情况。结果 成功构建了AE1缺失后 16位氨基酸的C末端表达质粒。结论 重组质粒pGADT7 AE1 c end truncation对酵母无毒性 ,不能激活检测基因 ,可作为酵母双杂交系统中的靶基因。

Objective To construct anion exchange 1 C terminal expression plasmid with Asp896～val911 deletion. Methods The last 16 amino acids (Asp896～Val911)of the AE1 c end were deleted and amplified by PCR from pFAST Bac AE1 c end.The AE1 c end truncation gene fragment was cloned into pGADT7 from AD domain in Yeast two hybrid system.The recombinant plasmid was transformed into Yeast AH109 and the expression in the Yeast was observed. Results Constructed the new vector pGADT7 AE1 c end truncation.Conclusion pGADT7 AE1 c end truncation has nontoxic to the yeast, which can serve as a target gene of Yeast two hybrid system.