| 表达PRRSV 强弱毒Nsp2的Marc 145细胞系的建立及Nsp2蛋白对PRRSV复制的影响 |
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| 引用本文： | 王凤雪,刘 准,温永俊,冷 雪,李真光,武 华.表达PRRSV 强弱毒Nsp2的Marc 145细胞系的建立及Nsp2蛋白对PRRSV复制的影响[J].西北农业学报,2012,21(3):1-6. |
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| 作者姓名： | 王凤雪 刘 准 温永俊 冷 雪 李真光 武 华 |
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| 作者单位： | 中国农业科学院特产研究所,吉林,132109 |
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| 基金项目： | 国家“十一五”科技支撑计划(2009BADB4B02);国家“863”计划(2011AA10A213) |
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| 摘 要： | 为探讨Nsp2蛋白对猪繁殖与呼吸综合征病毒(PRRSV)复制的影响,采用体外克隆法构建表达高致病性PRRSV TJ株和其致弱毒TJM株Nsp2基因的真核表达质粒pEGFP-TJ Nsp2和pEGFP-TJMNsp2,含有增强式绿色荧光蛋白表达盒,瞬时转染重组质粒到Marc-145细胞系单层,利用G418抗性筛选建立细胞系,通过PCR和RT-PCR鉴定。PRRSV感染筛选的细胞系,检测病毒TCID50。结果建立稳定表达TJ株和TJM株Nsp2基因的猴肾细胞系Marc-145-TJ Nsp2和Marc-145-TJM Nsp2;在表达PRRSV TJ株和TJM株Nsp2蛋白的Marc-145细胞上感染PRRSV病毒,在复制早期病毒时增殖速度快,即Nsp2蛋白对PRRSV复制早期有正向调控作用,并且强毒的Nsp2此作用更明显。
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| 关 键 词： | 猪繁殖与呼吸综合征病毒 Nsp2蛋白 细胞系 复制 |
The Construction of Marc 145 Cell Lines Expressing Nsp2 Gene of PRRSV and Nsp2 Protein Effect on PRRSV Replication |
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| WANG Fengxue,LIU Zhun,WEN Yongjun,LENG Xue,LI Zhenguang and WU Hua.The Construction of Marc 145 Cell Lines Expressing Nsp2 Gene of PRRSV and Nsp2 Protein Effect on PRRSV Replication[J].Acta Agriculturae Boreali-occidentalis Sinica,2012,21(3):1-6. |
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| Authors: | WANG Fengxue LIU Zhun WEN Yongjun LENG Xue LI Zhenguang and WU Hua |
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| Institution: | (Institute of Special Wild Economic Animal and Plant Science,Chinese Academy Agricultural Sciences,Jilin 132109,China) |
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| Abstract: | To investigate the effects of Nsp2 protein on porcine reproductive and respiratory syndrome virus replication,the Nsp2 gene of highly pathogenic PRRSV TJ and attenuated TJM were amplified with RT-PCR and cloned into the plasmid pEGFP-N1,which containing enhanced green fluorescent protein expression box.The constructed plasmid pEGFP-TJ Nsp2 and pEGFP-TJM Nsp2 were transfected into Marc-145 cells and screened by G418.Anti-G418 Marc-145-TJ Nsp2 and Marc-145-TJM Nsp2 cells were obtained and the expression of Nsp2 protein in anti-G418 Marc-145-TJ Nsp2 and Marc-145-TJM Nsp2 cells were proved by PCR,RT-PCR.The Marc-145-TJ Nsp2 and Marc-145-TJM Nsp2 cells were infected by PRRSV,and determined TCID50.Results showed the cells expressing Nsp2 gene of highly pathogenic PRRSV TJ and attenuated TJM were stable.PRRSV replication was fast in early stage on these cells.That is to say,Nsp2 protein plays a positive role in early phase of PRRSV proliferation,and the effect of Nsp2 protein of highly pathogenic PRRSV TJ was more obvious. |
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| Keywords: | Porcine reproductive and respiratory syndrome virus Nsp2 protein Cell lines Replication |
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