Objective: To explore the biological function of rhoptry protein 38 (ROP38) of Toxoplasma gondii, and to identify the reactogenicity of the recombinant protein (rROP38).
Methods: The ROP38 was amplified by RT-PCR from T. gondii RH strain, and was cloned into prokaryotic expression vector pET-28a (+). The recombinant plasmid was transformed into E. coli BL21 (DE3) competent cells. Then the rROP38 was analyzed by SDS-PAGE and identified by Western blot.
Results: SDS-PAGE showed that rROP38 was efficient expression with a molecular weight of about 43 kD. Western blot showed that rROP38 reacted with antibody of His tag or human positive antibody, which indicated that ROP38 had good reactogenicity and could be a serological diagnostic antigen.
Conclusions: The study successfully obtains the rROP38 of T. gondii with good reactogenicity.
[摘要]目的 原核表达弓形虫棒状体蛋白ROP38, 并对重组蛋白rROP38的反应原性进行鉴定。方法 根据已发表 的ROP38基因序列设计引物, 通过RT-PCR方法扩增弓形虫RH株的ROP38基因。将ROP38部分基因克隆到原核表达 载体pET-28a (＋) 后, 重组质粒转化大肠杆菌BL21 (DE3) 感受态细胞, IPTG诱导表达并优化表达条件, SDS-PAGE分析表 达情况, 并通过Western blot鉴定其反应原性。结果 SDS-PAGE显示在上清和包涵体均有rROP38表达, 分子量约为43 kD。Western blot结果显示, rROP38蛋白能被His标签抗体和感染弓形虫的人阳性血清识别, 说明ROP38具有良好的反 应原性, 可以作为潜在的血清学诊断抗原。结论 本研究成功获得了具有良好反应原性的弓形虫重组蛋白rROP38。.
Keywords: Parasitophorous vacuole (PV); Prokaryotic expression; Rhoptry protein 38 (ROP38); Toxoplasma gondii.