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当前位置: 首页 > 文章 > hPRRSV GD07株Nsp2基因真核表达载体的构建和表达 黑龙江畜牧兽医 2013 (11) 22-25
Position: Home > Articles > Construction and eukaryocyte expression of Nsp2 gene of highly pathogenic GD07 strain of PRRSV Heilongjiang Animal Science and Veterinary Medicine 2013 (11) 22-25

hPRRSV GD07株Nsp2基因真核表达载体的构建和表达

作  者：
牛晓芸;王艳丽;钟望;贺东生
单  位：
华南农业大学兽医学院;华南农业大学兽医学院广东省人畜共患病重点实验室广东大华农动物保健品股份有限公司
关键词：
猪繁殖与呼吸综合征(PRRS);非结构蛋白-2(Nsp2);真核表达
摘  要：
为了深入研究猪繁殖与呼吸综合征病毒(PRRSV)Nsp2蛋白的免疫特性、结构与功能,试验根据GenBank公布的PRRSV CH-1a株Nsp2基因的核苷酸序列设计1对特异性引物,PCR扩增目的基因,亚克隆至真核表达载体pcDNA3.1(+)中,构建重组质粒pcDNA3.1(+)-Nsp2,并对其进行经PCR、双酶切鉴定及DNA序列测定;将构建的重组质粒转染到Marc-145细胞中,用SDS-PAGE、Western-blot对表达产物进行鉴定。结果表明:RT-PCR扩增出的Nsp2基因大小与预期一致,经SDS-PAGE及Western-blot鉴定Nsp2蛋白在Marc-145细胞中成功表达。
译  名：
Construction and eukaryocyte expression of Nsp2 gene of highly pathogenic GD07 strain of PRRSV
作  者：
NIU Xiao-yun1,WANG Yan-li1,ZHONG Wang1,HE Dong-sheng1,2(1.College of Veterinary Medicine,South China Agricultural University,Guangzhou 510642,China;2.Guangdong Provincial Key Laboratory of Research and Application for Veterinary Biological Products Technology,Guangzhou 510640,China)
关键词：
porcine reproductive and respiratory syndrome(PRRS);non-structural protein 2(Nsp2);eukaryotic expression
摘  要：
To further the immunogenicity,structure and function of Nsp2 protein of porcine reproductive and respiratory syndrome virus(PRRSV),a pair of primers was designed according to the nucleotide sequence of the Nsp2 gene of the CH-1a strain of PRRSV published in the GenBank.The Nsp2 gene of hPRRSV GD07 strain was successfully amplified by RT-PCR,and subcloned into a eukaryotic expression vector pcDNA3.1(+) to establish the recombinant plasmid pcDNA3.1(+)-Nsp2,and then the recombinant eukaryotic expression plasmid was identified using PCR,double restriction enzyme digestion,and DNA sequence.The recombinant plasmid was transfected into the Marc-145 cells,and the expressed product was identified by SDS-PAGE and western blotting.The results showed that the size of Nsp2 gene was consistent with the expected.SDS-PAGE and western-blot showed that the protein of Nsp2 was successfully expressed in the Marc-145 cells.

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