Prokaryotic expression of Helicobacter pylori gene hp1188

HAN Fei; et al

doi:10.13406/j.cnki.cyxb.2010.06.002

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Prokaryotic expression of Helicobacter pylori gene hp1188

更新时间：2024-01-17

- Prokaryotic expression of Helicobacter pylori gene hp1188
- Journal of Chongqing Medical University Vol. 35, Issue 6, Pages: 822-825(2010)
- 作者机构：
  
  1. 重庆三峡中心医院微免科
  2. 重庆医科大学基础医学院:病原生物学教研室
  3. 神经科学研究中心4. 基础医学实验教学中心病原生物学与免疫学实验室
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.2010.06.002
  CLC： Q78
- Published：2010，
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[1]韩飞,杨致邦.幽门螺杆菌hp1188基因的原核表达[J].重庆医科大学学报,2010,35(06):822-825.

HAN Fei, et al. Prokaryotic expression of Helicobacter pylori gene hp1188. [J]. Journal of Chongqing Medical University 35(6):822-825(2010)
[1]韩飞,杨致邦.幽门螺杆菌hp1188基因的原核表达[J].重庆医科大学学报,2010,35(06):822-825. DOI： 10.13406/j.cnki.cyxb.2010.06.002.

HAN Fei, et al. Prokaryotic expression of Helicobacter pylori gene hp1188. [J]. Journal of Chongqing Medical University 35(6):822-825(2010) DOI： 10.13406/j.cnki.cyxb.2010.06.002.

摘要

目的:重组表达幽门螺杆菌（Helicobacter pylori

H.pylori）hp1188基因

为研究该基因所编码蛋白在H.pylori的黏附过程中的作用奠定基础。方法:以H.pylori标准株NCTC11637基因组DNA为模板

PCR扩增hp1188基因片段

克隆至质粒pQE-30中获得重组质粒pQE30-hp1188

经DNA测序确认后将重组质粒转化大肠杆菌DH5α

IPTG诱导表达。Ni2+-NTA树脂亲和层析纯化表达蛋白

SDS-PAGE检测表达

Westernblot鉴定相应抗原表位。结果:扩增的hp1188基因片段全长810bp

与基因文库中的hp1188基因同源性达98%。SDS-PAGE显示表达产物相对分子量约为30600Dalton

与预期相符;大肠杆菌裂解液中表达产物接近可溶性蛋白总量的一半

但沉淀中仍能检测到表达蛋白。目标蛋白纯化后均一性接近90%

Westernblot证实与多克隆抗体有良好结合能力。结论:成功克隆了hp1188基因

并在大肠杆菌DH5α中获得高效表达。

Abstract

Objective:To express the recombinant protein of the hp1188 gene of Helicobacter pylori NCTC 11637 in order to lay a foundation for the study on adhesion. Methods:The hp1188 gene was amplified from NCTC 11637 chromosomal DNA by PCR.The PCR product was inserted into the expression vector pQE-30.The recombinant vector pQE30-hp1188 was identified by DNA sequencing and transformed to E.coli DH5α for expression under IPTG induction.The recombinant protein was purified by Ni

-NTA agarose and identified by SDS-PAGE and western blot. Results:The sequence of hp1188 gene amplified was about 810 bp with a 98% sequence homology with hp1188 genes in GenBank. SDS-PAGE determined that the molecular weight of its expression products was 30 600 Dalton. The expression products of E.coli DH5α lyzate was about 50% of total soluble proteins

and the pellet also had expressed. Its purity was over 90% after purified with Ni

-NTA agarose and Western blot showed that the recombinant protein possesses fine binding capacity with the rabbit anti-H.pylori. Conclusion:hp1188 gene was successfully cloned and highly expressed in E.coli DH5α.

关键词

幽门螺杆菌hp1188基因表达

Keywords

Helicobacter pylorihp1188 geneexpression

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