肝胆胰外科杂志2012，Vol.24Issue(2)：123-127,5.
大鼠GSK3β基因RNAi腺病毒载体的构建与鉴定及其对肝卵圆细胞增殖的促进作用
Construction and identification of RNAi adenovirus vectors against GSK3β gene in rat and its promotion to the proliferation of hepatic oval cells
摘要
Abstract
bjective To construct and identify adenovirus vectors with the specific siRNA of rat GSK3β gene,andobservethoseeffectsontheproliferationofhepaticoval cellsWBF-344. Methods Two pairs of shRNA sequences against different parts of GSK3β gene were cloned into the RNAi plasmids vectors pGenesil-1. 1 by recombinant DNA technology to construct shRNA expression vectors pGenesil-l. 1-GSK3β. The vectors Pdc312 and recombinant plasmids were digested respectively,transformed and connected. Adenovirus vectors Pdc312-GSK3β were then built and amplified and identified with PCR and Western blot. Adenovirus vectors and backbone plasmids Ppe3-Fll were co-transfected to 293 ceils by liposome means. Adenovirus particles AdDC312-GSK3β were packaged and produced. The titer of adenovirus wasmeasured. After the viral supernatant infectedWB-F344 cells, fluorescent levels were observed under fluorescence microscope,and the expression of GSK3β was detected with Western blotting. The changes of cells proliferation were measuredwith CCK8. Results It was confirmed that RNAi plasmids pGenesil-1.1-GSK3β and RNAi adenovirus vectors AdDC312-GSK3βwere successfully constructed with PCR and Western blot. The adenovirus vectors could interfere the expression of GSK3 β in WBF-344 cells. The results of CCK8 showed that interferingGSK3β gene could promote the proliferation of WBF-344.Conclusion RNAi adenovirusvectorsagainst GSK3β genehave successfully constructed, andthey can steadily expressinthehepaticovalcellslineWBF-344,thuspromotethecells' proliferation.关键词
GSK3β/RNAi/腺病毒/肝卵圆细胞/大鼠Key words
CSK3β/ RNAi/ adenovirus/ hepatic oval cell/ rats分类
医药卫生引用本文复制引用
谢元康,钟骏桥,蒋磊,季晓克,郭旭,张启瑜,单云峰..大鼠GSK3β基因RNAi腺病毒载体的构建与鉴定及其对肝卵圆细胞增殖的促进作用[J].肝胆胰外科杂志,2012,24(2):123-127,5.基金项目
国家自然科学基金资助项目(30700800). （30700800）
