平榛主要过敏原一个片段区基因的克隆表达及免疫鉴定  

Cloning and expression of a fragment of Corylus heterophylla 1 and identifying its immunologic competence

作　　者：李娜[1,2] 胡东生[1,2] 刘志刚[2] 林斯里[2] 罗新萍[2] 黄钟[2] 

机构地区：[1]郑州大学公共卫生学院流行病与卫生统计学教研室,河南郑州450001 [2]深圳大学医学院,广东深圳518060

出　　处：《细胞与分子免疫学杂志》2011年第1期4-6,共3页Chinese Journal of Cellular and Molecular Immunology

基　　金：深港创新圈项目资助(200701);深圳大学团队基金资助(200904)

摘　　要：目的:克隆表达平榛(Corh)主要过敏原Corh1的一个片段区基因,并纯化表达的蛋白及检测其免疫学活性。方法:采用生物信息学方法选取Corh1的主要抗原表位区,设计带有酶切位点的特异性引物,采用RT-PCR方法扩增目的基因,将其导入pMD18-T载体中测序。将测序正确的质粒双酶切,并将获得的片段基因导入pET-32a中表达。通过Ni2+亲和层析柱纯化重组蛋白,采用Westernblot方法检测其IgE结合活性。结果:克隆并获得了Corh1的主要表位区基因,基因开放阅读框为243个碱基,编码81个氨基酸,蛋白相对分子质量(Mr)约为9000。表达的蛋白以可溶性为主,纯化出的蛋白有较好的免疫原性。结论:成功地克隆表达了Corh1的主要表位区基因,蛋白具有良好的免疫学活性。AIM： To clone, express, and identify a fragment of Corh1 from Corylus heterophylla, METHODS： Through bioinformatics predication, the antigenic epitope of Corh1 was selected. A fragment gene of Corh1 was amplified by PCR and cloned into pMD18T vector for sequencing. Then the fragment gene was sub cloned into pET-32a expression vector for expression, and then purified by metal （Ni2~） chelating affinity chromatography. The immunogenicity was tested by Western blot. RESULTS： The length of the fragment gene was 243 bp, coding 81 amino acids; the relative molecular mass of recombinant protein was 9 000. And the fragment of Corh1 was mainly expressed as soluble protein, purified protein has good immunogenicity. CONCLUSION： The fragment gene of Corh1 was successfully cloned and expressed in this study, and the recombinant protein possessed good IgEbinding capacity.

关 键 词：平榛主要过敏原 Corh1 抗原表位 克隆表达 

分 类 号：R392.8[医药卫生—免疫学]