斯氏副柔线虫免疫相关基因CSY原核表达载体的构建及生物信息学分析

Bioinformatic analysis and construction of its prokaryotic expression vector of immune-related gene CSY of Parabronema skrjabini

采用RT-PCR克隆斯氏副柔线虫CSY基因的CDS区,限制性内切酶对目的片段与pET30a（＋）原核表达载体进行双酶切,构建重组表达载体,利用生物信息学软件对该基因理化性质、结构特点进行分析。结果显示：CSY基因全长774bp,编码255个氨基酸,蛋白质理论大小值为29 240,理论等电点为7.53,蛋白质不稳定指数为40.09;跨膜结构和信号肽分析表明CSY蛋白无跨膜区,无信号肽区域。结果表明：成功构建了pET-30a（＋）-CSY重组表达载体;生物信息学分析表明CSY蛋白是一种抗原性较高的亲水性蛋白,可能有7个B细胞抗原表位和8个T细胞抗原表位,有望用于斯氏副柔线虫的免疫诊断抗原和疫苗候选抗原。

In order to construct prokaryotic expression vecto of pET30a（＋）-CSY, the secretory protein CSY from Parabronerna skrjabini was analyzed for its antigenic epitopes and gene by bioinformatics. The CDS region of Parabronema skrjabini CSY gene was obtained by RT-PCR. Restriction endonuclease was used to double digestion of the target fragment and pET30a（＋） pro- karyotic expression vector and then constructed the recombination expression vector. The physicochemical properties and structural feature of CSY gene were analyzed by bioinformatics software. The results indicated that CSY gene was 774 bp and encoding 255 amino acids. The protein weights is 29 240. Theoretical pI is 7.53. Instability index is 40.09. Transmernbrane structure and signal peptide analysis showed that CSY protein had no transmembrane domains and signal peptide region. The recombinant expression vector of pET-30a（＋）-CSY was successfully constructed in this study. CSY protein may have seven B-cell epitopes and eight T-cell epitopes. The CSY antigen has been promised as a candidate for immune diagnostics and a vaccine against Parabronerna skrjabini.