siRNA XIAP表达载体的构建及其生物学作用

目的构建靶XIAP的siRNA表达载体，研究其对Hep3B细胞中XIAP表达的影响。方法根据siRNA设计原则，设计并合成三段XIAP的siRNA序列，克隆到pRNAT—U6．1／Neo质粒构建重组质粒；经序列分析确认质粒构建成功后，将重组质粒转染入人肝癌细胞Hep3B，通过RT．PCR、Westernblot和免疫组化分析X!AP的表达。结果重组质粒pRNAT—u6．1／Neo．XIAP经过基因序列分析，siRNA片段成功插入，转染不同siRNAXIAP质粒均可使XIAP的翻译和蛋白表达水平不同程度的降低。结论成功构建并筛选出靶向XIAP基因的siRNA表达载体，为进一步研究XIAP在调控肝癌细胞凋亡方面的作用奠定了基础。

Construction of recombinant plasmid siRNA of XIAP and its biological role

Objective To construct siRNA expression vector of XIAP, and study its effect on XIAP expression in Hep3B cells. Methods Three XIAP siRNA sequences were designed, synthesized, and cloned to pRNAT -U6. 1/Neo. The successfully constructed recombinant plasmid was determined by se- quence analysis, and will be transfected into Hep3B. The best interference plasmid were analyzed by RT- PCR, Western blot, and immunohistochemistry. Results The plasmid of pRNAT-U6. 1/Neo- XIAP was constructed successfully, the trans-fected with different plasmid of siRNA XIAP can lower significantly XI- AP. Conclusions The siRNA vector of XIAP gene was constructed successfully. It will be a basis for the study of XIAP function in apoptosis regulation of the Hepatoma cells.