脑心肌炎病毒BD2株VP1基因的原核表达及生物信息学分析

发布时间：2018-05-20 17:31

  本文选题：脑心肌炎病毒 + VP基因　； 参考：《中国兽医学报》2017年04期

【摘要】：应用RT-PCR技术扩增脑心肌炎病毒(EMCV)BD2株VP1基因,将其克隆至原核表达载体pET-32a(+),利用在线分析软件对该基因所编码的蛋白序列进行生物信息学分析。将重组质粒pET-32a-VP1转化到E.coli BL21(DE3)感受态细胞中,在不同浓度的IPTG和不同温度条件下对目的蛋白进行诱导表达。结果显示,EMCV VP1基因全长831bp,编码277个氨基酸,VP1蛋白为酸性、亲水性蛋白质,蛋白空间结构以α-螺旋、β-折叠和无规则卷曲为主。经SDS-PAGE分析可知,VP1蛋白在37℃,1.0mmol/L IPTG的诱导条件下以包涵体形式获得了高效表达,重组蛋白的相对分子质量约为49 300。Western blot结果表明其具有良好的反应原性。综上所述,EMCV VP1蛋白是一种抗原性较高的亲水性蛋白,在原核系统中能高效表达,该研究为进一步建立相应的抗体检测方法和DNA疫苗的研制提供理论基础。
[Abstract]:The RT-PCR technique was used to amplify the VP1 gene of encephalitis virus strain EMCVV BD2, and cloned it into the prokaryotic expression vector pET-32a (pET-32a). The protein sequence encoded by the gene was analyzed by bioinformatics using on-line analysis software. The recombinant plasmid pET-32a-VP1 was transformed into E.coli BL21 (DE3) competent cells, and the target protein was induced to express under different concentrations of IPTG and different temperature. The results showed that the total length of EMCV VP1 gene was 831 BP, encoding 277 amino acid protein VP1 as acidic and hydrophilic protein. The spatial structure of the protein was 伪 -helix, 尾 -fold and irregular curl. SDS-PAGE analysis showed that the VP1 protein was highly expressed in the form of inclusion body under the induction of 1.0 mmol / L IPTG at 37 鈩，

本文编号：1915583