Random shuttle mutagenesis: Gonococcal mutants deficient in pilin antigenic variation

Ian J. Mehr, H. Steven Seifert*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

33 Scopus citations

Abstract

Shuttle mutagenesis has been adapted to randomly mutate the genome of Neisseria gonorrhoeae (gonococcus; Gc). A size-restricted plasmid library of Gc strain FA1090 was mutated with the mini-transposon mTnEGNS. Randomness was tested by checking for transposon insertion bias between vector and insert DNA, Gc transformation efficiency of individual mutated clones, and representation of unique clones before and after Gc transformation with a mutated pool of DNA. Mutants created by random shuttle mutagenesis were screened, using a colony-based polymerase chain reaction assay, for the ability to undergo pilin antigenic variation. Out of 8064 mutants screened, 22 unique transposon insertion mutants were found to be antigenic variation deficient (Avd). The Avd mutants were separated into five types according to recombination defect-associated phenotypes, including colony growth, natural DNA transformation competence, and repair of DNA damage caused by ultraviolet radiation.

Original languageEnglish (US)
Pages (from-to)1121-1131
Number of pages11
JournalMolecular Microbiology
Volume23
Issue number6
DOIs
StatePublished - 1997

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

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