摘要
A 0.5kb antisense RNA fragment was designed targeting the 5’ non-coding region (NCR), translation initiation site and potential transcriptional active region of the sole immediate early gene(IE180) of Pseudorabies virus(PRV). The antisense RNA fragment was amplified by PCR and inserted into the downstream of hCMV IE promoter/enhancer of the eukaryotic expression vector pCI-neo. The resulting antisense RNA expression plasmid(pCI-0.5) was transfected into PK-15 cells and the transfected cells were selected by G418.The expression of antisense RNA was confirmed by RT-PCR. In order to assess the antiviral potency of the cell lines expressing the antisense RNA, the antisense RNA cell lines and the control cell lines were infected with 0.1 pfu of PRV strain Ea per cell. The infected supernatant were collected at different time points post-infection(p.i.) and the TCID\-(50) were further determined. The data showed that the cell lines expressing the antisense RNA could markedly inhibit the replication of PRV. The viral titers in cells expressing the antisense RNA was 156-fold less than control cells at 40 hp i.
A 0.5kb antisense RNA fragment was designed targeting the 5’ non-coding region (NCR), translation initiation site and potential transcriptional active region of the sole immediate early gene(IE180) of Pseudorabies virus(PRV). The antisense RNA fragment was amplified by PCR and inserted into the downstream of hCMV IE promoter/enhancer of the eukaryotic expression vector pCI-neo. The resulting antisense RNA expression plasmid(pCI-0.5) was transfected into PK-15 cells and the transfected cells were selected by G418.The expression of antisense RNA was confirmed by RT-PCR. In order to assess the antiviral potency of the cell lines expressing the antisense RNA, the antisense RNA cell lines and the control cell lines were infected with 0.1 pfu of PRV strain Ea per cell. The infected supernatant were collected at different time points post-infection(p.i.) and the TCID\-(50) were further determined. The data showed that the cell lines expressing the antisense RNA could markedly inhibit the replication of PRV. The viral titers in cells expressing the antisense RNA was 156-fold less than control cells at 40 hp i.
出处
《中国病毒学》
CSCD
2005年第1期89-91,共3页
Virologica Sinica
基金
国际科学基金(IFS
B/3092 1)
湖北省自然科学基金(2000J100)
关键词
伪狂犬病毒
立即早期基因
反义RNA
抑制
Pseudorabies virus(PRV)
Immediate-early gene
Antisense RNA
Inhibition
