Comprehensive Identification of Conditionally Essential Genes in Mycobacteria Proc Natl Acad Sci U S A. 2001 Oct 23;98(22):12712-7. Epub 2001 Oct 16.

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Presentation transcript:

Comprehensive Identification of Conditionally Essential Genes in Mycobacteria Proc Natl Acad Sci U S A Oct 23;98(22): Epub 2001 Oct 16.

How to Determine the Functions of Genes? DNA microarray PCR based mutagenesis Signature tagged mutagenesis (STM) Genomic Analysis and Mapping By in vitro Transposition (GAMBIT)

Nature Reviews Genetics 7: (2006)

Signature Tagged Mutagenesis (STM) Nature Reviews Genetics 7: (2006)

Genomic Analysis and Mapping By in vitro Transposition (GAMBIT) Annual Review of Microbiology 53: (1999)

Nature Reviews Genetics 7: (2006)

Generation of Mutant Libraries

Labeled RNA Synthesis

Transposon-Site Hybridization Rich mediumMinimal medium

Minimal medium Rich medium

Rich medium Minimal medium Gene Ratio (Rich/Minimal) Auxotrophic Mutations Identified by TraSH

leuB leuD ilvA Superpathway of Leu, Val, and Ile biosynthesis BioCyc

Superpathway of sulfate assimilation and Cys biosynthesis cysD cysH BioCyc

EMBO Journal 15: (1996)

Codon Usage of M. bovis TAT/TACTyr2.06% (100% of Tyr) TTA/CTALeu0.63% (6.4% of Leu) ATAIle0.22% (5.2% of Ile) GTAVal0.48% (5.6% of Val) TAA/TAGStop0.14% (46.6% of Stop) TATAAT-10 element CBS

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