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Updated: Aug 14, 2026
Generation, Amplification, and Titration of Recombinant Respiratory Syncytial Viruses
Published on: April 4, 2019
[Construction and effect of the recombinant pshRNA plasmid against respiratory syncystial virus M2-1 gene]
Yu-xia Cui1, Juan Zhou, Ping Fang
1Immunology Laboratory, Children's Hospital, Chongqing University of Medical Science, Chongqing 400014, China.
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).
Conclusion:
The recombinant pshRNA7816 plasmid which targeted the mRNA of RSV M2-1 gene showed a significant and specific anti-RSV effect.
Insights
RNA interference targeting the M2-1 gene significantly inhibits respiratory syncytial virus (RSV) replication in cell cultures. This novel RNAi plasmid demonstrates specific antiviral effects, reducing viral load and improving cell survival.
Area of Science:
- Molecular Virology
- RNA Interference Therapeutics
- Antiviral Drug Discovery
Context:
- Respiratory syncytial virus (RSV) is a leading cause of infant respiratory infections globally.
- Current treatments for RSV lack efficacy, necessitating novel therapeutic strategies.
- RNA interference (RNAi) offers a gene-level approach to inhibit viral replication.
Purpose:
- To investigate the efficacy of RNA interference targeting the M2-1 gene of RSV.
- To construct and evaluate a recombinant RNAi plasmid (pshRNA7816) for anti-RSV activity.
- To assess the impact of M2-1 gene-specific RNAi on viral replication and cell viability in vitro.
Summary:
- A recombinant plasmid, pshRNA7816, targeting the RSV M2-1 gene mRNA was successfully constructed.
- Transfection of pshRNA7816 into Hep2 cells significantly reduced RSV-induced cytopathic effects and viral titer.
- Viral plaque assays and MTT assays confirmed a reduction in viral load and enhanced survival of RSV-infected cells.
Impact:
- The study demonstrates a significant and specific anti-RSV effect of the M2-1 gene-targeting RNAi plasmid.
- This finding supports the potential of RNAi technology as a therapeutic strategy against RSV infections.
- Further research may lead to the development of novel antiviral treatments for RSV.
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