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目的分析和预测猪带绦虫成虫延伸因子(elongation factor-1,EF-1)基因及其编码蛋白的结构域特性,并进行原核表达。方法利用在线生物信息学网站及工具进行序列分析、预测其编码的延伸因子的理化特性、抗原表位、翻译后的修饰、功能域、亚细胞定位、拓扑结构、二级结构、三维空间构象等,将其编码区序列克隆岛原核表达载体pET-28 a(+)上,测序鉴定重组质粒。结果该基因全长1 657 bp,编码区为186~1 533 bp,编码448个氨基酸,为全长基因;无跨膜区,具有多个磷酸化位点,蛋白的理化性质稳定,理论分子量为49 011.5 Da;没有质体、线粒体定位序列;十二烷基-聚丙烯酰胺凝胶电泳(SDS-PAGE)结果表明,目的基因在大肠埃希菌BL21~DE3中表达成功。结论发现猪带绦虫成虫延伸因子基因,成功构建重组原核表达质粒。
OBJECTIVE: To analyze and predict the elongation factor (EF-1) gene of adult Taenia solium and the characteristics of its encoded protein, and prokaryotic expression. Methods Using the online bioinformatics website and tools, sequence analysis was performed to predict the physicochemical properties, antigenic epitopes, posttranslational modifications, functional domains, subcellular localization, topological structure, secondary structure, three-dimensional conformation The coding region sequence was cloned into prokaryotic expression vector pET-28 a (+), and the recombinant plasmid was identified by sequencing. Results The gene was 1 657 bp in length with a coding region of 186-1 533 bp encoding 448 amino acids and was a full-length gene. It had no transmembrane region and multiple phosphorylation sites. The physicochemical properties of the protein were stable. The theoretical molecular weight was 49 011.5 Da. There was no plastid and mitochondrial localization sequence. The results of SDS-PAGE showed that the target gene was successfully expressed in Escherichia coli BL21-DE3. Conclusion The gene of elongation factor of adult Taenia solium was found and the recombinant prokaryotic expression plasmid was successfully constructed.