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目的探讨通过载体在体外培养的细胞中表达发夹状siRNAs对HBV基因表达的抑制作用。方法构建表达针对HBV S基因mRNA的发夹状siRNAs的质粒pSilencer 2.1-U6-S,并与ayw亚型HBV全基因组表达质粒共转染体外培养的HepG2215细胞,用半定量RT-PCR分析目标mRNA表达丰度的变化,用ELISA法观察HBsAg表达的变化。结果siRNA处理组细胞上清中HbsAg和HBeAg含量分别比空白对照组降低了63.4%和68.0%,细胞中的2.1kb的信使RNA降低了75.2%。结论siRNA在体外培养的细胞中能有效地抑制HBV基因的表达。
OBJECTIVE: To investigate the inhibitory effect of hairpin siRNAs on HBV gene expression in vitro by using vector. Methods Plasmid pSilencer 2.1-U6-S expressing hairpin siRNAs targeting HBV S gene mRNA was constructed and transfected into HepG2215 cells co-transfected with the ayw subtype HBV whole genome expression plasmids. Semi-quantitative RT-PCR was used to analyze target mRNA The changes of expression abundance were observed by ELISA method HBsAg expression changes. Results The contents of HbsAg and HBeAg in the supernatant of siRNA-treated cells decreased by 63.4% and 68.0% respectively compared with the blank control group, and the mRNA of 2.1kb in cells decreased by 75.2%. Conclusion siRNA can effectively inhibit the expression of HBV gene in cultured cells.