title: [Cloning and prokaryotic expression of the outer membrane protein gene PorB of Neisseria gonorrhoeae].
abstract: OBJECTIVE
  To construct a fused expression vector of the outer membrane protein gene PorB of Neisseria gonorrhoeae, express the fusion protein in the prokaryotic system, and obtain a gene recombination protein, for the purpose of preparing the ground for further research on the pathopoiesis and immune protective response of PorB.
  
  
  METHODS
  A pair of primers were designed according to the known sequence of the PorB gene, and the PorB gene was amplified by PCR from the genome of Neisseria gonorrhoeae 29403 and cloned into the prokaryotic expression plasmid pGEX-4T-1 to generate pGEX-4T-PorB recombinants. The recombinant plasmid pGEX4T-PorB was transferred into competent cells E. coli BL21. After confirmed by restriction endonuclease digestion, PCR and DNA sequencing analysis, the recombinant protein was induced to express by isopropyl-beta-D-thiogalactoside (IPTG), and examined by SDS-PAGE and Western blotting.
  
  
  RESULTS
  Restriction endonuclease digestion, PCR amplification and DNA sequencing analysis showed that the PorB gene of 1 047 bp was amplified from Neisseria gonorrhoeae DNA, and the recombinant plasmid pGEX-4T-PorB was successfully constructed and highly expressed in E. coli.
  
  
  CONCLUSIONS
  The prokaryotic expression vector of pGEX-4T-PorB was successfully constructed and efficiently expressed in the prokaryotic system, which has provided a basis for further study on the biological activity of the PorB protein, as well as animal immune experiment and detection of Neisseria gonorrhoeae, and its application as a mucosal immune vaccine.
authors.affiliation: Laboratory of Pathogenic Biology, School of Preclinical Medicine, Dali University, Dali, Yunnan 671000, China. wangyan1358@163.com
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.forename: Lei
authors.initials: L
authors.lastname: Zhang
authors.forename: Li
authors.initials: L
authors.lastname: Zhang
authors.forename: Han
authors.initials: H
authors.lastname: Wang
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D009344:Neisseria gonorrhoeae / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D018272:Porins / Q000235:genetics / Q000378:metabolism*
chemical_list: D001425:Bacterial Outer Membrane Proteins; D018272:Porins; C057699:porin protein, Neisseria