Checks and balances with use of the Keio collection for phenotype testing

Sandra J. Aedo, Helena R. Ma, Mark P. Brynildsen

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

The Keio single gene knockout collection comprises approximately 4000 mutants of E. coli K12 strain BW25113, where each mutant contains a kanamycin resistance cassette in place of a single nonessential gene. This mutant library has proven to be incredibly useful in the fields of bacteriology, chemical genomics, biotechnology, and systems biology, which is evidenced by the greater than 3800 citations that the article describing its construction has garnered in the approximate first 11 years since its publication. Among the various applications of the collection, the most extensive use has been in the assessment of how loss of specific gene function influences phenotypes. In this chapter, we describe pitfalls with use of the collection and procedures that can be employed to ensure robust phenotype assessment of mutations in the library. These include procedures for thorough confirmation of gene deletions by PCR, phage transduction of mutated loci to new host strains, and strategies for genetic complementation.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages125-138
Number of pages14
DOIs
StatePublished - Jan 1 2019

Publication series

NameMethods in Molecular Biology
Volume1927
ISSN (Print)1064-3745

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Genetics

Keywords

  • Escherichia coli
  • Gene duplication
  • Knockout
  • P1 phage transduction

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    Aedo, S. J., Ma, H. R., & Brynildsen, M. P. (2019). Checks and balances with use of the Keio collection for phenotype testing. In Methods in Molecular Biology (pp. 125-138). (Methods in Molecular Biology; Vol. 1927). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-9142-6_9