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目的:研究用RNA干涉技术(RNAi)稳定抑制PC-1基因表达对前列腺癌细胞C4-2生物学行为的影响。方法:用DNA重组技术构建携带与短的发夹结构的干涉RNA相对应DNA序列的重组质粒pSC394-414,采用脂质体将pSC394-414重组质粒稳定转染C4-2细胞,PCR分析外源DNA序列的整合情况,RTPCR及Western印迹鉴定RNAi抑制PC1基因表达的细胞株,MTT试验分析该细胞的生长速度,软琼脂克隆形成实验检测其非锚着依赖性生长能力。结果:获得了用RNAi稳定抑制PC-1基因表达的细胞株C4-2-pSC394-414,该细胞株中的PC-1mRNA及PC1蛋白的表达均显著降低,细胞生长速度显著减慢,软琼脂克隆形成能力显著下降。结论:抑制PC-1基因表达,可降低C4-2细胞生长速度及非锚着依赖性生长能力。
Objective: To investigate the effect of RNA interference (RNAi) on the inhibition of PC-1 gene expression on the biological behavior of prostate cancer cell line C4-2. METHODS: Recombinant plasmid pSC394-414 carrying the DNA sequence corresponding to the interfering RNA with short hairpin structure was constructed by DNA recombination technique. The recombinant plasmid pSC394-414 was stably transfected into C4-2 cells by lipofectamine. The expression of exogenous DNA sequence integration, RTPCR and Western blotting were used to identify the cell lines that RNAi inhibited the expression of PC1 gene. The cell growth rate was analyzed by MTT assay and the non-anchorage-dependent growth ability by soft agar colony formation assay. Results: The cell line C4-2-pSC394-414 which stably inhibited the expression of PC-1 gene by RNAi was obtained. The expression of PC-1 mRNA and PC1 protein in the cell line was significantly decreased, the cell growth rate was significantly slowed, Clonal formation capacity decreased significantly. Conclusion: Inhibition of PC-1 gene expression can reduce the growth rate and non-anchorage-dependent growth of C4-2 cells.