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| 猪繁殖与呼吸综合征病毒ORF7基因原核表达载体的构建及序列分析 | |
| 引用本文： | 张永富,马国文,刘月焕,刘永宏,李明刚,张秋雨,刘锋.猪繁殖与呼吸综合征病毒ORF7基因原核表达载体的构建及序列分析[J].中国动物检疫,2008,25(12):25-27,30. |
| 作者姓名： | 张永富 马国文 刘月焕 刘永宏 李明刚 张秋雨 刘锋 |
| 作者单位： | 1. 内蒙古民族大学动物科技学院,内蒙古通辽,028000;北京市农林科学院畜牧兽医研究所,北京,100089;北京庄笛浩禾生物医学科技有限公司,北京,100043 2. 内蒙古民族大学动物科技学院,内蒙古通辽,028000 3. 北京市农林科学院畜牧兽医研究所,北京,100089 4. 北京市农林科学院畜牧兽医研究所,北京,100089;内蒙古农业大学动物科学与医学学院,呼和浩特,010018 5. 北京庄笛浩禾生物医学科技有限公司,北京,100043 |
| 摘 要： | 根据GenBank中美洲型猪繁殖与呼吸综合征病毒(PRRSV)ORF7基因序列,设计合成一对引物,应用RT-PCR方法扩增出猪繁殖与呼吸综合征病毒(PRRSV)的ORF7基因(N基因)。将所扩增片段克隆入原核表达载体pET-32a(+),pET-ORF7重组质粒转化DH5a宿主菌后,经双酶切、PCR鉴定后挑选阳性克隆测序鉴定并对插入的ORF7基因序列进行分析。结果表明,ORF7基因的原核表达载体构建成功。ORF7基因序列与美洲型的ORF7基因核苷酸同源性为92.8%～99.7%,与LV株的相应基因核苷酸同源性为65.3%;推导的氨基酸与美洲型相应基因的同源性为92.0%～99.2%,与LV株的同源性为65.3%,系统进化树表明该PRRSV属于美洲型。本研究为N蛋白的进一步研究和制备诊断性抗原奠定了基础。 |
| 关 键 词： | 猪繁殖与呼吸综合征病毒 ORF7基因 原核表达载体 |
Construction and Sequence analysis of the Prokaryotic Expression Vector for ORF7 Gene of Porcine Reproductive and Respiratory Syndrome Virus |
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| ZHANG Yongfu,MA Guowen,LIU Yuehuan,LIU Yonghong,LI Minggang,ZHANG Qiuyu,LIU Feng.Construction and Sequence analysis of the Prokaryotic Expression Vector for ORF7 Gene of Porcine Reproductive and Respiratory Syndrome Virus[J].China Journal Of Animal Quarantine,2008,25(12):25-27,30. | |
| Authors: | ZHANG Yongfu MA Guowen LIU Yuehuan LIU Yonghong LI Minggang ZHANG Qiuyu LIU Feng |
| Institution: | 1.College of Animal Science and Technology,Inner Mongolia University For the Nationalities,Tongliao Inner Mongolia 028000;2. Institute of Animal Husbandry and Veterinary Medicine, Beijing Academy of Agriculture and Forestry Sciences, Beijing 100089;3.Beijing DIHO Biomedicai Science &Technology Co.,Ltd, Beijing 100043;4. College of Animal Science and Medicine, Inner Mongolia Agriculture University,Huhhot Inner Mongolia 010018 ) |
| Abstract: | A pair of primers were designed and synthesized according to the ORF7 gene sequences of American genotype porcine reproductive and respiratory syndrome virus (PRRSV) in GenBank.The PRRSV ORF7 gene (N gene) was amplified by RT-PCR.The amplified gene fragment was then cloned into the prokaryotic expression vector pET -32a (+ ),the recombinant plasmid was transformed into host cell DH5a ,the selected positive clones were subjected to sequence analysis of the inserted ORF7 gene after double enzyme digestion and PCR identification.The result showed that the construction of the prokaryotic expression vector for ORF7 gene was successful.The nucleotide homology between the cloned ORF7 gene and North American genotype ORF7 gene was92.8% ～99.7% and the nucleotide homology between the cloned ORF7 gene and the LV genotype ORF7 gene was 65.3 %. The deduced amino acids of the cloned ORF7 gene showed 92.0%～99.2%and 65.3%homology to strains North American and LV. The phylogenetic trees revealed the strain was clustered within North American genotype.The study provides a base for further research of N protein and production of diagnosis antigen. |
| Keywords: | porcine reproductive and respiratory syndrome virus ORF7 gene prokaryotic expression vector |
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