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RNA干扰是一种具有序列特异性的基因沉默,能够触发具有相应序列的mRNA的降解.构建具有双靶点的RNAi质粒表达载体,与单靶点表达载体比较,探讨其对结肠癌细胞增殖的抑制作用.本研究分别构建了针对Bcl-2、C-Raf和Bcl-2/C-Raf靶基因的质粒表达载体,通过LipofectamineTM2000介导转染人结肠癌细胞系HCT-8后,检测相应转染组靶基因的mRNA和蛋白质表达量,测定各组细胞活性,研究RNAi对各组癌细胞增殖的抑制率.结果表明,分别转染3种质粒表达载体后,3组结肠癌细胞中相应靶基因的mRNA和蛋白质表达量均降低;转染双靶点干扰质粒的试验组;其细胞活性低于单靶点组;对于针对Bcl-2,C-Raf和Bc-l2/C-Raf基因的3组干扰实验,RNAi对结肠癌细胞增殖的抑制率分别为43.87%,40.64%和63.85%.RNAi是结肠癌细胞中的一种功能途径,以质粒作为表达载体,同时具备Bc-l2/C-Raf双靶点的表达载体,对结肠癌细胞增殖的抑制作用要明显优于单靶点表达载体,双靶点质粒表达载体在结肠癌的基因治疗中是有潜力的.
RNAi is a kind of sequence-specific gene silencing that can trigger the degradation of mRNA with the corresponding sequence.We constructed a RNAi plasmid vector with two targets and compared with the single-target expression vector to explore its effect on the proliferation of colon cancer cells The purpose of this study was to construct a plasmid expression vector targeting Bcl-2, C-Raf and Bcl-2 / C-Raf target genes, respectively, and then transfected into human colon cancer cell line HCT-8 by LipofectamineTM2000, The mRNA and protein expression levels of the target genes were determined and the activity of each group was determined.The inhibitory rate of RNAi on the proliferation of each group was studied.The results showed that after transfection of three kinds of plasmid expression vector, The mRNA and protein expression levels of Bcl-2, C-Raf and Bc-l2 / C-Raf genes were lower in the experimental group transfected with the double-target interfering plasmid than in the single target group 3 groups of interference experiments, RNAi on colon cancer cell proliferation inhibition rates were 43.87%, 40.64% and 63.85% .RNAi is a functional pathway in colon cancer cells to plasmid as an expression vector, with Bc-l2 / C -Raf dual target expression vector for colon cancer cell proliferation The inhibitory effect is obviously better than the single target expression vector, and the dual target plasmid expression vector has potential in the gene therapy of colon cancer.