癌胚抗原相关细胞黏附分子6-shRNA及双自杀基因yCDglyTK的联合基因载体构建

Construction of combined gene vector of carcinoembryonic antigen associated cell adhesion molecule 6(CEACAM6)-shRNA and double suicide gene of yCDglyTK

目的构建新型靶向联合双自杀基因载体pCDNA3.1-yCDglyTK-CEACAM6-shRNA。方法根据已知的癌胚抗原相关细胞黏附分子6(Carcinoembryonic antigen associated cell adhesion molecule 6,CEACAM6)序列构建含有hU6启动子和CEACAM6-shRNA表达框的重组质粒,并将CEACAM6-shRNA表达框(含hU6启动子)通过双酶切方式亚克隆至双自杀基因载体pCDNA3.1-yCDglyTK,构建新型靶向联合双自杀基因载体pCDNA3.1-yCDglyTK-CEACAM6-shRNA。通过酶切、测序等鉴定重组质粒。结果实验所构建的最终载体酶切产物所得的片段大小为7.5 kb,与预期片段一致。测序证实最终载体与pCDNA3.1-yCDg-lyTK-CEACAM6-shRNA序列一致。结论成功构建新型靶向联合双自杀基因载体pCDNA3.1-yCDg-lyTK-CEACAM6-shRNA。

【Objective】 To construct a newly target-combined double suicide gene vector pCDNA3.1-yCDglyTKCEACAM6-shRNA.【Methods】 Constructed a recombinant plasmid which contains hU6 promoter and CEACAM6shRNA frame by known sequence of CEACAM6（Carcinoembryonic antigen associated cell adhesion molecule 6）.Then subcloned the CEACAM6-shRNA frame which contains hU6 promoter to the double suicide gene vector pCDNA3.1-yCDglyTK by double enzyme cutting,and the newly target-combined double suicide gene vector pCDNA3.1-yCDglyTK-CEACAM6-shRNA was constructed.The newly recombinant plasmid was identified by enzyme cutting and sequencing.【Results】 The size of the newly constructed vector fragment by enzyme cutting was 7.5kb,which is in line with anticipation.The result of sequencing showed that the newly constructed target-combined double suicide gene vector was in accord with pCDNA3.1-yCDglyTK-CEACAM6-shRNA.【Conclusion】 The newly target-combined double suicide gene vector pCDNA3.1-yCDglyTK-CEACAM6-shRNA is constructed successfully.