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目的为从基因水平抑制呼吸道合胞病毒(respiratory syncytial virus,RSV)的复制,针对RSV的M2-2基因构建小发卡结构状RNA(short hairpin RNA,shRNA)重组质粒,在细胞水平观察shRNA对RSV复制的影响。方法将成功构建的针对RSV M2-2基因的重组质粒pshRNA8260转染HEp-2细胞,利用光镜观察pshRNA8260对RSV致HEp-2细胞病变效应(cytopathogenic effect,CPE)的影响并计算CPE抑制率,空斑形成实验检测RSV滴度变化。结果成功构建了针对人RSV M2-2基因mRNA的pshRNA8260重组质粒,研究发现pshRNA8260能明显改善RSV所致的病变效应,降低RSV在细胞内复制的病毒滴度。结论针对RSV M2-2基因的pshRNA8260重组质粒具有明显的特异性抗RSV效应的作用。
The purpose is to inhibit the replication of respiratory syncytial virus (RSV) at the gene level and to construct a short hairpin RNA (shRNA) recombinant plasmid against the M2-2 gene of RSV. The effect of shRNA on RSV The effect of copy. Methods The recombinant plasmid pshRNA8260 against RSV M2-2 gene was transfected into HEp-2 cells. The effect of pshRNA8260 on the cytopathogenic effect (CPE) of HEp-2 cells induced by RSV was observed by light microscopy. The CPE inhibition rate, Plaque formation assay was used to detect changes in RSV titer. Results The pshRNA8260 recombinant plasmid targeting human M2 gene was successfully constructed. The results showed that pshRNA8260 can significantly improve the pathogenic effect induced by RSV and reduce the virus titer in RSV replication in cells. Conclusion The pshRNA8260 recombinant plasmid targeting RSV M2-2 gene has obvious anti-RSV effect.