Construction of prokaryotic expression plasmid of mtrC protein ofNeisseria gonorrhoeae and its expression inE. coli

Abstract

In order to provide a rational research basis for detection of resistance ofNeisseria gonorrhoeae transferable resistance (mtr) efflux system, plasmid pET-28a(+) encoding mtrC gene was constructed and the related target protein was expressed inEscherichia coli (E. coli) DE3. The fragments of mtrC gene ofNeisseria gonorrhoeae from the standard strains were amplified and cloned into prokaryotic expression plasmid pET-28a(+) with restriction endonuclease to construct recombinant pET-mtrC which was verified by restriction endonuclease and DNA sequencing. The recombinant was transformed intoE. coli DE3 to express the protein mtrC induced by IPTG. The results showed mtrC DNA fragment was proved correct through restriction endonuclease and DNA sequencing. Its sequence was 99.5% homologus to that published on GeneBank (U14993). A 48. 5 kD fusion protein which was induced by IPTG was detected by SDS-PAGE. It was concluded that the construction of prokaryotic expression plasmid of mtrC protein ofNeisseria gonorrhoeae was correct and the fusion protein was successively expressed inE. coli.

Keywords

Neisseria gonorrhoeae / prokaryotic expression / multiple transferable resistance efflux system

References

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