家蚕过敏原Profilin的表达、纯化、免疫学鉴定及其生物信息学分析

发布时间：2019-01-25 20:53

【摘要】：目的:克隆表达家蚕(Bombyx mori)Profilin蛋白,鉴定其免疫原性并进行B细胞抗原表位预测和构建分子进化树。方法:从NCBI上获取家蚕Profilin蛋白的基因序列,合成该基因并构建到pet-28a表达载体上,重组质粒转化至E.coli BL21,IPTG诱导基因表达,通过亲和层析获取高纯度蛋白;用Western blot方法对重组蛋白进行过敏原性鉴定;通过DNAStar软件分析其潜在的B细胞抗原表位;应用MEGA5.05进行序列比对,并构建分子进化树。结果:成功表达出重组蛋白,且重组蛋白与家蚕过敏患者血清有一定的Ig E结合。结论:成功克隆表达出具有免疫原性的Profilin蛋白,并成功预测出其B细胞抗原表位和构建出分子进化树。
[Abstract]:Objective: to clone and express (Bombyx mori) Profilin protein of Bombyx mori, identify its immunogenicity, predict the epitope of B cell antigen and construct a molecular evolutionary tree. Methods: the gene sequence of Profilin protein of Bombyx mori (Bombyx mori) was obtained from NCBI. The gene was synthesized and constructed into pet-28a expression vector. The recombinant plasmid was transformed into E.coli BL21,IPTG induced gene expression, and high purity protein was obtained by affinity chromatography. Western blot method was used to identify the allergen of recombinant protein, DNAStar software was used to analyze the potential B cell epitopes, MEGA5.05 was used to sequence alignment, and molecular evolutionary tree was constructed. Results: the recombinant protein was successfully expressed, and the recombinant protein had certain Ig E binding to the sera of silkworm allergy patients. Conclusion: Profilin protein with immunogenicity was cloned and expressed successfully, and its B cell epitopes were successfully predicted and molecular evolutionary tree was constructed.
【作者单位】：深圳大学过敏反应与免疫学研究所;北京协和医院变态反应科;
【基金】：国家卫生部公益性行业科研专项(No.2015SQ00136) 广东省工程技术研究开发中心项目(No.2013158925) 广东省对外科技合作项目(No.2013B051000088) 深圳市科技计划基础研究项目(No.JCYJ20140828163633991、JCYJ20140418095735604)
【分类号】：R392

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