New cloning vectors to facilitate quick allelic exchange in gram-negative bacteria

Adrian Mejia-Santana, Cameron J. Lloyd, Karl E. Klose

Research output: Contribution to journalArticlepeer-review

Abstract

New cloning vectors have been developed with features to enhance quick allelic exchange in gram-negative bacteria. The conditionally replicative R6K and transfer origins facilitate conjugation and chromosomal integration into a variety of bacterial species, whereas the sacB gene provides counterselection for allelic exchange. The vectors have incorporated the lacZ alpha fragment with an enhanced multicloning site for easy blue/white screening and priming sites identified for efficient in vivo assembly or other DNA assembly cloning techniques. Different antibiotic resistance markers allow versatility for use with different bacteria, and transformation into an Escherichia coli strain capable of conjugation enables a quick method for allelic exchange. As a proof of principle, the authors used these vectors to inactivate genes in Vibrio cholerae and Salmonella typhimurium.

Original languageEnglish (US)
Pages (from-to)117-120
Number of pages4
JournalBioTechniques
Volume70
Issue number2
DOIs
StatePublished - Jan 15 2021

Keywords

  • Cloning
  • Genetic engineering
  • Gram-negative
  • Microbiology

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry, Genetics and Molecular Biology(all)

Fingerprint Dive into the research topics of 'New cloning vectors to facilitate quick allelic exchange in gram-negative bacteria'. Together they form a unique fingerprint.

Cite this