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目的:在原核系统中表达并纯化人类组氨酰TRNA合成酶类似物蛋白,制备多克隆抗体。方法:设计引物扩增其开放阅读框,重组入原核表达载体PQE30,并在大肠杆菌M15中诱导表达;应用纯化的重组蛋白免疫新西兰大白兔。结果:①成功构建人类组氨酰TRNA合成酶类似物的原核表达载体;②经NI亲和层析获得纯度较高的重组蛋白;③制备了高滴度的兔抗人类组氨酰TRNA合成酶类似物蛋白的多克隆抗体。结论:原核表达载体的构建、重组蛋白的表达、纯化及多克隆抗体的制备为今后研究该蛋白的功能提供了良好的基础。
OBJECTIVE: To express and purify the human histamine TRNA synthetase analogue protein in prokaryotic system to prepare polyclonal antibody. Methods: The primers were designed to amplify the open reading frame (ORF), then inserted into prokaryotic expression vector PQE30 and induced in E. coli M15. The purified recombinant protein was used to immunize New Zealand white rabbits. Results: ①The prokaryotic expression vector of human histamine TRNA synthetase was constructed successfully; ② The recombinant protein with high purity was obtained by NI affinity chromatography; ③ High titer rabbit anti-human histidyl TRNA synthase Polyclonal antibodies to analog proteins. Conclusion: The construction of prokaryotic expression vector, expression of recombinant protein, purification and preparation of polyclonal antibody provide a good foundation for the future study of the function of this protein.