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中国农学通报

中国农学通报 ›› 2005, Vol. 21 ›› Issue (8): 13-13.

所属专题： 畜牧兽医

• 目次 • 上一篇    下一篇

反义RNA介导的抗猪传染性胃肠炎病毒感染

于晓龙   

  • 出版日期:2005-08-05 发布日期:2005-08-05

Antisense RNA-Medidiated Resistance to TGEV Infection In IBRS2 Cells

Yu Xiaolong   

  • Online:2005-08-05 Published:2005-08-05

摘要： 构建了重组逆转录病毒表达质粒plxas-N，该质粒能表达互补猪传染性胃肠炎病毒N基因全长的反义RNA序列。用质脂体方法将重组质粒plxas-N转染至PA317细胞中，经抗生素G418（500μg/ml）筛选出稳定的产毒细胞克隆。分别扩大培养细胞克隆，取其上清液感染小鼠成纤维细胞NIH3T3，测定细胞克隆产生的重组病毒滴度，并用高滴度重组病毒感染IBRS2细胞。提取被感染的IBRS2细胞的总RNA，RT-PCR证明plxas-N整合到IBRS2细胞基因组。通过TGEV分别感染IBRS2和IBRS2抗性细胞产生的病变，结果表明该反义RNA对病毒的复制有抑制作用，其抑制率近50%。

Abstract: A recombinant expression plasmid, plxas-N, was designed to produce antisense RNA targeted to the N gene full length of Transmissible gastroenteritis virus of swine, TGEV. The plxas-N was transfected into packaging cells line PA317 with Lipofectamine. The viral supenatants of the clones selected with G418 (500μg /ml) were detected by murine fibre cell NIH3T3.The highest viral titer pseudovirus was used to infect IBRS2 cell. Total cellular RNA from the IBRS2 cell, and RT-PCR analysis indicated that the plxas-N had inserted into the genome of IBRS2 cell. Antiviral activitity of the antisense RNA was evaluated by cytopathic effect using cultured IBRS2 cell and anti- IBRS2 cell infected by TGEV respectively, and the inhibitory rate was about 50 %.