Identifying essential genes in Mycobacterium tuberculosis by global phenotypic profiling

Methods Mol Biol. 2015;1279:79-95. doi: 10.1007/978-1-4939-2398-4_6.


Transposon sequencing (TnSeq) is a next-generation deep sequencing-based method to quantitatively assess the composition of complex mutant transposon libraries after pressure from selection. Although this method can be used for any organism in which transposon mutagenesis is possible, this chapter describes its use in Mycobacterium tuberculosis. More specifically, the methods for generating complex libraries through transposon mutagenesis, design of selective pressure, extraction of genomic DNA, amplification and quantification of transposon insertions through next-generation deep sequencing are covered. Determining gene essentiality and statistical analysis on data collected are also discussed.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Bayes Theorem
  • Chromosomes, Bacterial / genetics
  • DNA Transposable Elements / genetics
  • DNA, Bacterial / genetics
  • Gene Library
  • Genes, Essential*
  • Genomics / methods*
  • Markov Chains
  • Models, Statistical
  • Molecular Sequence Data
  • Mutagenesis, Insertional / genetics
  • Mycobacterium tuberculosis / genetics*
  • Phenotype


  • DNA Transposable Elements
  • DNA, Bacterial