应用载体介导的RNAi技术抑制胰腺癌细胞K-RAS^(Asn12)的表达  

Inhibition of K-RAS^(Asn12) Expression by Vector-based RNA Interference in Human Pancreatic Cancer Cell Line

作　　者：孟繁杰[1] 付泽娴[1] 张峰[1] 李保东[1] 谢绍建[1] 蔡建辉[1] 

机构地区：[1]河北医科大学第二医院外科外科研究中心,石家庄050000

出　　处：《中国生物工程杂志》2006年第4期86-90,共5页China Biotechnology

基　　金：国家自然科学基金资助项目(30140008);河北省自然科学基金资助项目(302511)

摘　　要：为了研究载体介导的RNAi(RNA interference)技术特异地抑制K-RASAsn12(K-RAS第12位密码子突变形式为GAT)在胰腺癌细胞中的表达,合成两条编码针对K-RASAsn12突变特异性短发夹 RNA(short hairpin RNA,shRNA)序列的单链DNA,并将其克隆到pSilenCircle载体中,构建含目的基因片段的重组质粒pSC-K-RASAsn12。同法构建含GFP基因片断的重组质粒pSC-GFP做为对照。用其分别瞬时转染人胰腺癌AsPC-1细胞和BxPC-3细胞后,经半定量RT-PCR和Western blot检测K- RAS的表达水平。结果表明构建的针对K-RASAsn12的编码突变特异性shRNA的质粒表达载体可特异的抑制胰腺癌细胞的K-RASAsn12表达,但对野生型的K-RAS(K-RASWT)表达无影响。To silence the expression of K-RASAsn12 in human pancreatic cancer cell line by vector-based RNAi（RNA interference） technique, two single-strand DNA sequences encoding mutant-specific shRNA （short haipin RNA） for K-RASAsn12 were synthesized and then inserted into pSilenCircle. The recombinant plasmid was called pSC-K-RASAsn12. According to the same method, pSC-GFP encoding shRNA for GFP was gained. Both recombinant plasmids were transfected into human pacreatic cancer cell line AsPC-1 and BxPC-3. The expression level of K-RASAsn12 was detected by semi-quantitative RT-PCR and Western blot. The result indicated that the recombinant plasmid edcoding mutant-specific shRNA for K-RASAsn12 can inhibit significantly the expression of K- RASAsn12, without affection of wild-type K-RAS（K-RASWT） in Human Pancreatic Cancer Cell Line.

关 键 词：胰腺癌K-RAS RNAi短发夹RNA 

分 类 号：Q752[生物学—分子生物学]