[Construction and effect of the recombinant pshRNA plasmid against respiratory syncystial virus M2-1 gene].
OBJECTIVE
Respiratory syncystial virus (RSV) is the most common cause of lower respiratory infections in infants worldwide. There is no reliable vaccine or antiviral drug against RSV at present. RNA interference (RNAi) technology is a potent method to degrade expression of the cognate mRNA. In order to inhibit the replication of RSV at gene level, the effects of specific RNAi against M2-1 gene of RSV on inhibition of viral replication in cell culture system was observed in this study.


METHODS
RSV M2-1 gene, which plays a key role in RSV transcription, was chosen in this study and was used as target gene and recombinant plasmid pshRNA7816 targeting the mRNA of RSV M2-1 gene coding sequence was constructed. The pshRNA7816 was transfected into Hep2 cells. The effects of the pshRNA7816 on changes of cytopathogenic effect (CPE) of Hep2 cell induced by RSV infection were observed microscopically. Viral plaque forming assay and MTT assay were used to detect the viral titer change and protective function of the pshRNA7816 on RSV infected Hep2 cell.


RESULTS
The recombinant RNAi plasmid pshRNA7816 which targets the mRNA of RSV M2-1 gene was successfully constructed. The pshRNA7816 significantly reduced CPE of RSV infected Hep2 cells, reduced the viral titer of RSV in the cells (P < 0.001). The pshRNA7816 raised the survival rate of RSV infected Hep2 cells (P < 0.001). Non-specific pshRNA plasmid did not show anti-RSV effects (P > 0.05).


CONCLUSIONS
The recombinant pshRNA7816 plasmid which targeted the mRNA of RSV M2-1 gene showed a significant and specific anti-RSV effect.
Cui
Yu-xia
YX
Immunology Laboratory, Children's Hospital, Chongqing University of Medical Science, Chongqing 400014, China.
Zhou
Juan
J
Fang
Ping
P
Jiang
Li-ping
LP
Wang
Li-jia
LJ
Yang
Xi-qiang
XQ
D056945:Hep G2 Cells; D006801:Humans; D010957:Plasmids / Q000096:biosynthesis; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000096:biosynthesis; D012367:RNA, Viral / Q000235:genetics; D022261:Respiratory Syncytial Virus Vaccines / Q000096:biosynthesis*; D018113:Respiratory Syncytial Virus, Human / Q000187:drug effects* / Q000502:physiology; D014764:Viral Proteins / Q000235:genetics*; D014779:Virus Replication / Q000187:drug effects*
C507355:M2-2 protein, respiratory syncytial virus; D034741:RNA, Small Interfering; D012367:RNA, Viral; D022261:Respiratory Syncytial Virus Vaccines; D014764:Viral Proteins