Inhibition of Viral Repl ication by the Antisense RNA Targeting the 5′Regionsof Immediate2early Gene of Pseudorabies Virus

FANG Liu-rong; XIAO Shao-bo; YU Xiao-lan; XU Jian-xiang; CHEN Huan-chun*

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February 2005

FANG Liu-rong, XIAO Shao-bo, YU Xiao-lan, XU Jian-xiang and CHEN Huan-chun*. Inhibition of Viral Repl ication by the Antisense RNA Targeting the 5′Regionsof Immediate2early Gene of Pseudorabies Virus[J]. Virologica Sinica, 2005, 20(1): 89-91.

Citation: FANG Liu-rong, XIAO Shao-bo, YU Xiao-lan, XU Jian-xiang, CHEN Huan-chun*. Inhibition of Viral Repl ication by the Antisense RNA Targeting the 5′Regionsof Immediate2early Gene of Pseudorabies Virus .VIROLOGICA SINICA, 2005, 20(1) : 89-91.

伪狂犬病毒立即早期基因5′端特异性反义RNA对病毒增殖的影响

1.
华中农业大学动物医学院湖北武汉

摘要

伪狂犬病毒(Pseudorabies virus,PRV)属疱疹病毒科α 疱疹病毒亚科,基因组为线状双链 DNA,长约150kb,至少可编码 70~100 种蛋白质。根据伪狂犬病毒感染细胞后 DNA 转录、表达时间的先后可将PRV基因分为立即早期基因(Immediate-early gene),早期基因 (Early gene)和晚期基因...
- 伪狂犬病毒
- , 立即早期基因
- , 反义RNA
- , 抑制

Inhibition of Viral Repl ication by the Antisense RNA Targeting the 5′Regionsof Immediate2early Gene of Pseudorabies Virus

1.
College of Veterinary Medicine , Huazhong Agricultural University , Wuhan 430070 ,China

Abstract

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 vira ˎ̥,Verdana,Arial">titers in ˎ̥,Verdana,Arial; mso-font-kerning: 1.0pt; mso-bidi-font-family: 'Times New Roman'; mso-fareast-font-family: 宋体; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA">cells expressing the antisense RNA was 1562 fold less than control cells at 40 hp i.
- Pseudorabies virus（PRV）
- , Immediate-early gene
- , Antisense RNA
- , Inhibition

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Inhibition of Viral Repl ication by the Antisense RNA Targeting the 5′Regionsof Immediate2early Gene of Pseudorabies Virus

1. College of Veterinary Medicine , Huazhong Agricultural University , Wuhan 430070 ,China

Abstract: 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 vira ˎ̥,Verdana,Arial">titers in ˎ̥,Verdana,Arial; mso-font-kerning: 1.0pt; mso-bidi-font-family: 'Times New Roman'; mso-fareast-font-family: 宋体; mso-ansi-language: EN-US; mso-fareast-language: ZH-CN; mso-bidi-language: AR-SA">cells expressing the antisense RNA was 1562 fold less than control cells at 40 hp i.