花生过敏原Ara h 1蛋白的原核表达及致敏性研究

食品科学 ›› 0, Vol. ›› Issue (): 0-0.

• 生物工程 • 下一篇

花生过敏原Ara h 1蛋白的原核表达及致敏性研究

史云凤,张彤,陈沁

上海大学

收稿日期:2018-02-13 修回日期:2019-01-23 出版日期:2019-03-25 发布日期:2019-04-02
通讯作者: 陈沁 E-mail:chenqincc@staff.shu.edu.cn
基金资助:
国家自然科学基金

Prokaryotic expression of peanut allergen Ara h 1 and its allergenicity

Yunfeng Shi¹, ¹,qin chen

Received:2018-02-13 Revised:2019-01-23 Online:2019-03-25 Published:2019-04-02
Contact: qin chen E-mail:chenqincc@staff.shu.edu.cn

摘要/Abstract

摘要： 摘要：本实验从花生中提取RNA，用反转录PCR得到花生过敏Ara h 1基因，构建pET-28a-Ara h 1表达载体，转入Rosetta (DE3)中诱导，用镍离子亲和层析法纯化得到目的蛋白。SDS-PAGE结果显示目的蛋白分子量约为75kDa，与预计相符，经质谱鉴定为Ara h 1蛋白。用BALB/c小鼠模型和RBL细胞模型评价重组Ara h 1蛋白的致敏性。结果显示，重组Ara h 1蛋白致敏小鼠的空肠和肺组织发生病变，血清中特异性抗体、TH2型细胞因子、组胺含量升高，表明重组Ara h 1蛋白可以导致小鼠发生TH2型过敏反应；同时重组Ara h 1蛋白还导致RBL细胞脱颗粒，释放β-己糖苷酶，表明重组Ara h 1蛋白具有致敏性。

关键词: Ara h 1过敏原, 重组蛋白, 致敏性, BALB/c小鼠, RBL细胞

Abstract: Abstract: Ara h 1 gene was synthesized from total peanut RNA using oligo primers by reverse PCR. The Ara h 1 gene was cloned into pET-28a vector to construct a recombinant expression vector. Then the recombinant vector was transferred into the bacterial expression host rosetta (DE3) for inducible expression, and the target protein was purified by Ni affinity chromatography. SDS-PAGE results showed that the molecular weight of the expressed protein was about 75 kDa, which matched with the theoretical value. Mass spectrometric results confirmed that the target protein was Ara h 1.The allergenicity of recombinant Ara h 1 protein was evaluated by animal model of BALB/c mice and cell model of RBL cell. Inflammation occurred in the lung and jejunum, and the content of specific antibody, TH2 type cytokines and histamine were increased. Meanwhile, the recombinant Ara h 1 could trigger degranulation of RBL cell, and then released β- hexosaminidase，which indicated that recombinant Ara h 1 could cause TH2 type allergic reaction.

Key words: Key words: Ara h 1 allergen, recombinant protein, allergenicity, BALB/c mice, RBL cell

中图分类号:

Q789

史云凤张彤陈沁. 花生过敏原Ara h 1蛋白的原核表达及致敏性研究[J]. 食品科学, 0, (): 0-0.

Yunfeng Shi qin chen. Prokaryotic expression of peanut allergen Ara h 1 and its allergenicity[J]. FOOD SCIENCE, 0, (): 0-0.

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花生过敏原Ara h 1蛋白的原核表达及致敏性研究

Prokaryotic expression of peanut allergen Ara h 1 and its allergenicity

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