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当前位置: 首页 > 文章 > 牛病毒性腹泻-黏膜病病毒E_2基因在卡介苗中的优化表达 畜牧兽医学报 2014 (1) 94-100
Position: Home > Articles > Cloning and Optimizing Expression of E_2 Gene of Bovine Viral Diarrhea-mucosal Disease Virus in BCG Chinese Journal of Animal and Veterinary Sciences 2014 (1) 94-100

牛病毒性腹泻-黏膜病病毒E_2基因在卡介苗中的优化表达

作  者：
曾范利;张云;张梦;时坤;李建明;刘菲;李晶;杜锐
单  位：
教育部动物生产及产品质量安全重点实验室;吉林农业大学研究生学院;吉林省药用动物二级实验室;吉林农业大学动物科学技术学院;长春金赛药业有限责任公司;吉林农业大学中药材学院
关键词：
牛病毒性腹泻-黏膜病病毒;E_2基因;卡介苗;优化表达
摘  要：
为构建牛病毒性腹泻-黏膜病病毒(BVDV)基因工程活载体疫苗,根据已知的BVDV Changehun184毒株E2基因序列,利用DN.AStar生物学软件对其进行分析,预测了可能存在的抗原表位,设计6对引物,分别扩增了包含预测抗原位点的6个片段,1—297 aa、1—345 aa、1—374 aa、45—297 aa、45—345 aa和45—374 aa,克隆至穿梭表达载体pMV361中,经酶切、PCR扩增和测序证实已正确插入到表达载体中,构建了6个原核表达质粒。阳性重组质粒通过电转化到卡介苗(BCG)感受态中,热诱导后,进行SDS-PAGE分析和免疫印迹检测。结果表明6个重组质粒在卡介苗中均有不同程度的表达,表达产物与理论推测的相对分子质量一致。Western blot结果显示其融合蛋白能被牛抗BVDV的阳性血清识别,具有相应的反应原性。本研究为进一步研究BVDV基因工程活载体疫苗奠定了基础。
译  名：
Cloning and Optimizing Expression of E_2 Gene of Bovine Viral Diarrhea-mucosal Disease Virus in BCG
作  者：
ZENG Fan-li;ZHANG Yun;ZHANG Meng;SHI Kun;LI Jian-ming;LIU Fei;LI Jing;DU Rui;College of Chinese Medicine Materials,Jilin Agricultural University;College of Animal Science & Technology,Jilin Agricultural University;Changchun Kinsey Pharmaceutical Co.Ltd;College of Animal Science & Technology,J ilin Agricultural University;Key Laboratory of Animal Production,Product Quality and Security of Ministry of Education of the people's Republic of China,Level 2 Laboratory of Medical Animal of Jilin Province,College of Graduate,Jilin Agricultural University;
关键词：
Bovine viral diarrhea-mucosal disease virus(BVDV);;E_2 gene;;BCG;;optimizing expression
摘  要：
To develop genetic engineering living-vector vaccine of Bovine viral diarrhea-mucosal disease virus(BVDV),E_2 gene of BVDV Changchunl84 strain was selected and the complete sequence was analyzed,then the epitopes of E_2 protein were predicted by bioinformatics software.Six pairs of primers were designed according to E_2 gene sequence,and the epitopes coding regions(1-297,1-345,1-374,45-297,45-345,45-374) of E_2 gene were amplified by PCR and cloned into pMV361 vector.PCR,restriction endonuclease digestion and sequence analysis proved that the 6fragments were inserted into the expected position.It was indicated that the six prokaryotic expression plasmids were constructed.The positive recombinant plasmids were electro-transformed into BCG for expression at 45℃.The SDS-PAGE and Western blot results indicated that the recombinant proteins could react with polyclonal antibody against BVDV.Our study provides a basis for the further studies on genetic engineering living-vector vaccine of BVDV.

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