【摘要】 目的:构建针对HPV16 E6基因的RNA干扰表达载体,并研究其对宫颈癌HPV16 E6基因的抑制作用。方法:针对HPV16 E6基因序列设计shRNA(short hairpin RNAs,shRNAs)片断,构建针对HPV16 E6基因的RNA干扰(RNA interference,RN Ai)质粒表达载体,脂质体法转染宫颈癌Caski细胞株,应用荧光定量PCR及流式细胞术检测其对HPV16 E6 mRNA及蛋白表达的影响。结果:经PCR及测序证实3种表达载体构建成功,细胞试验表明3种表达载体均抑制了HPV16 E6的mRNA及蛋白的表达,其中CaskiB细胞HPV16E6mRNA的抑制率为89.5%,蛋白抑制率达98.1%。结论:RNAi表达载体可以有效的抑制HPV16 E6基因的表达。更多还原

【Abstract】 Objective:To obtain recombinant HPV16 E6 specific RNA interference expressi-on vectors and investigate the in- hibitory effects of the vectors on the expression of human papil-omavirus E6 gene.Methods:According to the computer aided design, 56nt oligonucleotide fragments containing different HPV16 E6 specific sequences were syntbsized and cloned into the expression vec- tors,The recombinants were transfected into cervical cancer cell /ine,Caski,with liposomes.Expression of E6 was detected by FQ- PCR(fluorescence quantitative PCR)and flow cytometry.Results:HPV16 E6 specific siRNA expression vectors were confirmed by PCR analysis and DNA sequencing.Three kinds of expression vectors could reduce the expressions of E6 mRNA and protein all in Caski-B cell.The inhibition ratio of E6 mRNA reduced to 89.5%,and protein inhibition ratio reached 98.1%.Conclusion:The RNAi expression vectors can effectively inhibit the expression of HPVE6 Gene.更多还原