In vitro recombination and identification of mutated fragment corresponding to regulation region of mtrR gene of Neisseria Gonorrhoeae

Abstract

A site-directed mutant DNA fragment was synthesized and transfected into clinical Neisseria Gonorrhoeae (NG) stains to construct the transformants that contained the corresponding mutagenesis of regulation region of mtrR gene. According to the technique of gene splicing by overlap extension (SOEing), a DNA segment with specific mutagenesis was constructed by two-step polymerase chain reaction (PCR). The mutation fragments EF could be used for the next experiment in which the mutation NG strains were induced. By comparing the recombinant EF fragments to the corresponding DNA fragments of clinical NG strains, 2 of these were not compatible completely. The results of sequencing revealed that there was a 9 bp deletion between the 45 to 54 inverted repeat sequence localized within the mtrR promoter. It can be confirmed that the fragments EF are the specifically designed mutant fragments.

Keywords

Nisseria gonorrhoeae / genes mutation / mtr drug resistance

References

[1] KaylaE. H., PanW. B., ShaferW. M., et al.. Resistance of Neisseria gonorrhea to antimicrobial hydrophobic agents is modulated by mtrRCDE efflux system. Microbiology, 1995, 141: 611-622

[2] PanW. H., LiaoW. Q., LiZ. W.. Effect of chromosome mtrR gene mutation on Neisseria gonorrhea multidrug resistance. J Chin Dermatol (Chinese), 1999, 32(3): 152-154

[3] HortonR. M.. PCR-mediated recombination and mutagenesis. SOEing together tailor-made genes. Mol Biotechnol, 1995, 32: 93-99

[4] HortonR. M.. In vitro recombination and mutagenesis of DNA. SOEing together tailor-made genes. Mol Biotechnol, 1997, 67: 141-149