| PRRSV陕西分离株ORF5基因克隆、序列分析及原核表达 |
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| 引用本文： | 王志昇,许信刚,童德文,邢福珊,陈 曦,唐麒麟,宁蓬勃.PRRSV陕西分离株ORF5基因克隆、序列分析及原核表达[J].西北农业学报,2010,19(8):1-6. |
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| 作者姓名： | 王志昇 许信刚 童德文 邢福珊 陈 曦 唐麒麟 宁蓬勃 |
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| 作者单位： | 西北农林科技大学,动物医学院,陕西杨凌,712100 |
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| 基金项目： | 陕西省科技攻关项目，国家大学生创新性实验计划项目，教育部新世纪优秀人才支持计划 |
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| 摘 要： | 克隆猪繁殖与呼吸综合征病毒(PRRSV)ORF5基因,并进行序列分析及原核表达。根据GenBank公布的PRRSV ORF5基因的核苷酸序列,设计并合成一对特异性引物,用RT-PCR方法扩增PRRSV陕西分离株糖基化囊膜蛋白基因ORF5,将其克隆入pGEM-T载体中,进行测序及序列分析。再设计另外1对引物扩增ORF5缺失编码N端31个氨基酸残基的基因片段,将截短的ORF5基因亚克隆入原核表达载体pET-32a中,在大肠杆菌BL21中进行原核表达。结果扩增到603 bp的PRRSV全长ORF5基因,序列分析表明,分离株与北美型代表株VR-2332和欧洲型代表株LV氨基酸同源性分别为87.8%和54.5%。因此推测陕西分离株属于北美型。SDS-PAGE可检测到大小约为38 ku的融合蛋白,主要以可溶性蛋白形式存在。West-ern-blot分析表明,重组蛋白可被PRRSV阳性血清所识别。
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| 关 键 词： | 猪繁殖与呼吸综合征病毒 陕西分离株 序列分析 原核表达 |
Cloning, Sequence Analysis and Prokaryotic Expression of ORF5 Gene of PRRSV Shaanxi Strain |
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| WANG Zhisheng,XU Xingang,TONG Dewen,XING Fushan,CHEN Xi,TANG Qilin and NING Pengbo.Cloning, Sequence Analysis and Prokaryotic Expression of ORF5 Gene of PRRSV Shaanxi Strain[J].Acta Agriculturae Boreali-occidentalis Sinica,2010,19(8):1-6. |
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| Authors: | WANG Zhisheng XU Xingang TONG Dewen XING Fushan CHEN Xi TANG Qilin and NING Pengbo |
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| Institution: | WANG Zhisheng,XU Xingang,TONG Dewen,XING Fushan,CHEN Xi,TANG Qilin and NING Pengbo (College of Veterinary Medicine,Northwest A&F University,Yangling Shaanxi 712100,China) |
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| Abstract: | According to the published complete nucleotide sequence of PRRSV(porcine reproductive and respiratory syndrome virus)in GenBank,a pair of primers specific to the full-length of ORF5 gene encoding the envelope glycoprotein GP5 was designed.The ORF5 gene fragment was amplified by RT-PCR and cloned into pGEM-T vector.The recombinant plasmid was sequenced and ORF5 gene was compared with GeneBank's other PRRSV strains.The ORF5 gene segment lacking the first 31 residues at the amino terminal region was amplified ... |
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| Keywords: | ORF5 |
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