斯氏副柔线虫SHR基因表达载体的构建及生物信息学分析

Bioinformatic analysis of Parabronema skrjabini SHR gene and construction of its prokaryotic expression vector

为探讨斯氏副柔线虫免疫相关SHR基因作为疫苗候选抗原及早期诊断抗原的可能性,提取斯氏副柔线虫的总RNA,用RT-PCR技术扩增SHR基因,并将PCR产物克隆到pMD19-T载体,构建原核表达载体pET-30a(+)-SHR,利用在线软件对该基因编码蛋白序列进行生物信息学分析。结果表明:克隆获得SHR基因全长1 083 bp,编码360个氨基酸,理论等电点为6.09,无信号肽和跨膜区;磷酸化预测含有29个磷酸化位点;结构分析发现,α-螺旋构成二级结构的主要成分,亲水性氨基酸比例超过60%;抗原表位预测表明,SHR蛋白是一种抗原性较高的亲水性蛋白。推测SHR有望用作斯氏副柔线虫的免疫诊断抗原和疫苗候选抗原。

The possibility of SHR gene as a vaccine candidate antigen and early diagnostic antigen was preliminary discussed in this research. The total RNA was extracted from Parabronema skrjabini and SHR gene was cloned by RTPCR. The PCR product was cloned into pMD19-T and a prokaryotic expression vector pET-30a (+)-SHR was then constructed. The protein structure and antigenic epitopes of SHR gene were analyzed by using online bioinformatics software. The results indicated that SHR gene was 1 083 bp in length,encoding 360 aminoacids and its theoretical pl is 6.09. Transmembrane structure and signal peptide analysis showed that the SHR protein contained a type of hydrophilic no transmembrane domains and signal peptide region and 29 phosphorylation sites. The secondary structure of SHR was mainly composed of alpha helix and the proportion of hydrophilic amino was over 60%. The antigenic epitopes prediction results indicated that SHR protein was a hydrophilic protein with high antigenicity. These results suggested that the SHR antigen has strong potential to be a candidate for immune diagnostics and used as a vaccine against P. skrjabini.