Identification of essential genes and synthetic lethal gene combinations in Escherichia coli K-12

Methods Mol Biol. 2015:1279:45-65. doi: 10.1007/978-1-4939-2398-4_4.

Abstract

Here we describe the systematic identification of single genes and gene pairs, whose knockout causes lethality in Escherichia coli K-12. During construction of precise single-gene knockout library of E. coli K-12, we identified 328 essential gene candidates for growth in complex (LB) medium. Upon establishment of the Keio single-gene deletion library, we undertook the development of the ASKA single-gene deletion library carrying a different antibiotic resistance. In addition, we developed tools for identification of synthetic lethal gene combinations by systematic construction of double-gene knockout mutants. We introduce these methods herein.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Conjugation, Genetic
  • DNA Primers / metabolism
  • Electroporation
  • Escherichia coli K12 / genetics*
  • Gene Knockout Techniques
  • Genes, Bacterial*
  • Genes, Essential*
  • Genes, Lethal*
  • Genes, Synthetic*
  • Genetic Complementation Test
  • Genomics / methods*
  • Molecular Sequence Data
  • Mutation
  • Polymerase Chain Reaction
  • Reproducibility of Results
  • Transformation, Genetic

Substances

  • DNA Primers