G250基因siRNA表达载体的构建与鉴定  

Construction and identification of G250 siRNA expression vector

作　　者：赵俊峰[1] 郑少斌[1] 姜耀东[1] 赵善超[1] 张香梅[1] 

机构地区：[1]南方医科大学南方医院,广东广州510515

出　　处：《山东医药》2008年第6期23-25,共3页Shandong Medical Journal

基　　金：广东省医学科研基金资助项目(B2006102)

摘　　要：目的构建肾癌相关抗原G250小分子干扰RNA(siRNA)表达载体,通过抑制G250基因的表达,探索肾癌基因治疗的新途径。方法根据基因库上的肾癌相关抗原G250 mRNA序列,设计并合成两端含有酶切位点的75个碱基的寡核苷酸链。寡核苷酸链退火后用T4DNA连接酶连接至线性化的质粒pRNAT-U6.1/Neo中,构建成重组质粒(命名为pshRNA-G250 a,pshRNA-G250b),进行酶切及序列鉴定。结果pshRNA-G250表达载体克隆构建成功,插入片段测序结果与合成的siRNA序列一致。结论成功构建pshRNA-G250表达载体,为进一步的研究及探索肾癌基因治疗奠定了基础。Objective To construct G250 siRNA expression vector in attempt to explore the new method of renal cell carcinoma gene therapy by suppressing G250 gene expression. Methods According to G250 mRNA sequence in the Genebank, a pair of 75-nt oligonucleotides, each containing the sites of restriction endonuclease at both ends, were designed and synthesized. Oligonucleotides were annealed and ligated with linearized pRNAT-U6. 1/Neo by T4DNA ligase. The recombinants （named pshRNA-G250a,pshRNA-G250b） were finally sequenced and identified by enzyme cutting and sequencing. Result~ pshRNA-G250 expression vector was successfully constructed and identified by double endonudease digestion. Sequence analysis of inserted fragment revealed the same sequence as synthesized siRNA oligonucleotides. Conclusions pshRNA-G250 expression vector has been successfully constructed, which will facilitate further studies of G250 function and its application in the treatment of renal cell carcinoma.

关 键 词：肿瘤相关抗原 G250基因 干扰RNA 遗传载体 基因疗法 肾肿瘤 

分 类 号：R737.11[医药卫生—肿瘤]