| 牛病毒性腹泻黏膜病毒E2基因的原核表达与免疫原性分析 |
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| 引用本文： | 王宇婷,马莉莉,李晓月,王士霞,毕莹,倪宏波.牛病毒性腹泻黏膜病毒E2基因的原核表达与免疫原性分析[J].安徽农业科学,2017,45(12). |
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| 作者姓名： | 王宇婷 马莉莉 李晓月 王士霞 毕莹 倪宏波 |
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| 作者单位： | 黑龙江八一农垦大学,动物科技学院,黑龙江大庆 163319;黑龙江八一农垦大学,动物科技学院,黑龙江大庆 163319;黑龙江八一农垦大学,动物科技学院,黑龙江大庆 163319;黑龙江八一农垦大学,动物科技学院,黑龙江大庆 163319;黑龙江八一农垦大学,动物科技学院,黑龙江大庆 163319;黑龙江八一农垦大学,动物科技学院,黑龙江大庆 163319 |
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| 基金项目： | 黑龙江省农垦总局"十二五"重点科技攻关项目 |
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| 摘 要： | 目的]原核表达牛病毒性腹泻黏膜病毒(BVDV)E2基因编码蛋白。方法]采用PCR方法从BVDV中扩增E2基因片段,与原核表达载体pET-32a连接,构建重组表达质粒pET-32a-E2,转化E.coli(Rosetta)感受态细胞,重组菌用1 mmol/L IPTG诱导表达E2蛋白,进行SDS-PAGE电泳,并用Ni-NTA亲和层析柱纯化目的蛋白,经Western blot分析鉴定免疫原性。结果]重组质粒pET-32aE2经PCR及酶切鉴定证明构建正确,重组质粒能够在大肠杆菌中大量表达,表达产物的分子质量大小约为58 kDa,纯化后E2重组蛋白浓度0.521 mg/mL,Western blot分析表明,其能被BVDV阳性血清识别,具有很好的免疫原性。结论]E2蛋白成功表达,为后续建立BVDV检测方法奠定了基础。
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| 关 键 词： | 牛病毒性腹泻黏膜病毒 E2基因 原核表达 |
Prokaryotic Expression and Inmunogenicity of E2 Gene of BVDV |
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| WANG Yu-ting,MA Li-li,LI Xiao-yue,WANG Shi-xia,BI Ying,NI Hong-bo.Prokaryotic Expression and Inmunogenicity of E2 Gene of BVDV[J].Journal of Anhui Agricultural Sciences,2017,45(12). |
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| Authors: | WANG Yu-ting MA Li-li LI Xiao-yue WANG Shi-xia BI Ying NI Hong-bo |
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| Abstract: | Objective]To prokaryotic express the bovine viral diarrhea-muscosal disease viruses E2 gene encoding protein.Method]Bovine viral diarrhea-muscosal disease viruses E2 gene were amplified by PCR and linked into the pET-32a prokaryotic expression vector,prokaryotic expression recombinant plasmid pET32a-E2 was constructed,which was then trandformed into E.coli(Rosetta)cells for protein expression.E2 protein of recombinant strains was induced to express by 1 mmol/L IPTG and SDS-PAGE electrophoresis,target protein was purified by Ni-NTA affinity chromatography column and the immunogenicity was identified by Western blot analysis.Result]The recombinant plasmid pET32a-E2 was confirmed by PCR,restriction enzyme.It had high-level expression in E.coli.SDS-PAGE showed that recombinant protein with molecular weight of 58 kDa,with concerntration of 0.521 mg/mL.Western blot showed that recombinant protien can reacts with positive serum,indicating good immunogenicity.Conclusion]E2 protein is expressed in successfully,which lays foundation for establishing BVDV detection method in future. |
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| Keywords: | Bovine viral diarrhea-mucosal disease viruses E2 gene Prokaryotic expression |
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