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目的:体外制备弓形虫醛缩酶(aldolase)蛋白及其多克隆抗体。方法:以弓形虫cDNA链为模板,PCR扩增醛缩酶基因,构建aldolase/pET30a原核表达系统;IPTG诱导表达aldolase-His6蛋白;用纯化的蛋白加免疫佐剂免疫SD大鼠,收集抗血清,制备多克隆抗体,亲和层析纯化并分析抗体的效价。结果:构建了aldolase原核表达系统,表达并纯化了aldolase-His6蛋白;获得了抗该蛋白的大鼠源性抗血清,纯化后的多克隆抗体效价为1∶4000。结论:在体外制备并纯化了aldolase-His6蛋白及其多克隆抗体,为后续研究aldolase的功能奠定了基础。
Objective: To prepare toxoplasma aldolase protein and its polyclonal antibody in vitro. Methods: Toxoplasma gondii cDNA chain was used as a template to amplify the aldolase gene by PCR. The prokaryotic expression system of aldolase / pET30a was constructed. The aldolase-His6 protein was induced by IPTG. SD rat was immunized with purified protein plus adjuvant. , Polyclonal antibodies were prepared, affinity purified and analyzed for antibody titer. Results: The aldolase prokaryotic expression system was constructed and the aldolase-His6 protein was expressed and purified. The rat-derived antiserum against the protein was obtained. The titer of the purified polyclonal antibody was 1: 4000. Conclusion: The aldolase-His6 protein and its polyclonal antibody were prepared and purified in vitro, which laid the foundation for the further study on the function of aldolase.