胰腺癌Bx-PC3细胞Survivin基因ShRNA质粒表达载体的构建  

Construction of eukaryotic expression vector expressing double shRNA sections targeting Survivin gene in Bx-PC3 cells

作　　者：黄鹤[1] 吴佩[1] 洪书剑[1] 茆家定[1] 芮景[1] 

机构地区：[1]皖南医学院附属弋矶山医院普外科

出　　处：《中国临床药理学与治疗学》2006年第6期655-658,共4页Chinese Journal of Clinical Pharmacology and Therapeutics

摘　　要：目的:构建针对胰腺癌Bx-PC3细胞Survivin基因2条shRNA的质粒表达载体。方法:以Survivin基因为靶点,通过自行设计并构建包含两段短发夹状RNA(small hairpin RNA,shRNA),携带有各自的U6启动子和终止码、共同的绿色荧光蛋白(greenfluorescent protein,EGFP)基因和Neo基因的pGene-sil-1SurvivinshRNA载体,利用脂质体介导的方法转染人胰腺癌Bx-PC3细胞,RT-PCR观察其对Bx-PC3细胞Survivin基因表达的影响。结果:HE1质粒和HE2质粒的酶切鉴定正确,质粒转化菌液HE1、HE2进行测序分析均为插入正确的克隆质粒。胰腺癌Bx-PC3细胞转染成功,转染细胞后SurvivinmRNA的表达有明显抑制。结论:实验构建针对胰腺癌细胞Survivin基因2条shRNA的质粒表达载体成功。AIM： To construct eukaryotic expression vector expressing double shRNA sections targeting Survivin gene. METHODS： Eukaryotic expression vector expressing double shRNA sections targeting Survivin gene were designed and chemically synthesized. They were directionally inserted into plasmid pGenesil-1 with respectively U6 promoter and termination code, the common green fluorescence protein （EGFP） gene and Neo gene. In this way, the vector of pGenesil-1 shRNA containing 2 sections of Survivin shRNA were constructed and they were transfected into the pancreatic cancer cell Bx-PC3. Transfection was detected by fluorescence microscope.The inhibition expression of Survivin mRNA was measured by RT-PCR. RESULTS： HE1 and HE2 plasmids were identified by the biocatalyst cut which confirmed the exactitude and were analyzed by the sequence analysis which verified the perfect clone plasmid inserted by them. CONCLUSION： A eukaryotic expression vector of double short hairpin RNA for Survivin gene is successfully constructed. The pancreatic cancer cells Bx-PC3 succeed to be transfected and expression of Survivin mRNA is inhibited obviously.

关 键 词：SURVIVIN基因 RNA干扰 短发夹RNA 胰 腺癌细胞 

分 类 号：R735.9[医药卫生—肿瘤]