Cloning,expression and immunogenicity analysis of helicobacter pylori oipA gene fragments
- VernacularTitle:幽门螺杆菌oipA基因片段的克隆表达及抗原性分析
- Author:
Feifei SHE
;
Ni LI
;
Xu LIN
;
Hao CHEN
- Publication Type:Journal Article
- Keywords:
Helicobacter pylori;
oipA gene fragment;
Clone;
Expression;
Antigenicity
- From:
Chinese Journal of Immunology
1986;0(04):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To express Helicobacter pylori oipA gene with different fragments and determine the protein and its peptides with finest antigenicity.Methods:oipA gene was obtained from international standard Hp NCTC 11637 and cloned into vector pGEM-T then was identified by PCR and sequencing.Six different fragments of oipA gene were amplified by PCR with the template pGEM-T/oipA.The gene fragments and expressing vector pET-42a were ligated.The recombinant vector were transformed into BL-21.The correct recombinants were identified by PCR,enzyme digestion and sequencing.Recombination protein and the peptides were detected by Western Breeze chemiluminescent after induced by IPTG.Optimization of the time and IPTG concentration for induction were conducted.The expressed products were affinity purified by Ni-NTA of His?Bind.The antigenicity of the recombination protein was determined with Western blot indicated by goat anti-Hp polyclonal antibody,and the recombination protein with finest antigenicity was selected with indirect ELISA.Results:The six oipA gene fragments could all expressed and the products were recognized by goat-anti Hp polyclonal antibody.The finest peptide with antigenicity was the smallest peptide.Conclusion:OipA gene fragments could be expressed in prokaryotic system and the smallest peptide had the finest antigenicity.It may be used as the reagent to detect corresponding antibody in serum.
