Comprehensive Identification of Conditionally Essential Genes in Mycobacteria

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Comprehensive Identification of Conditionally
Essential Genes in Mycobacteria
Proc Natl Acad Sci U S A. 2001 Oct
2398(22)12712-7. Epub 2001 Oct 16.
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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)

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Nature Reviews Genetics 7929-939 (2006)
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Signature Tagged Mutagenesis (STM)
Nature Reviews Genetics 7929-939 (2006)
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Genomic Analysis and Mapping By in vitro
Transposition (GAMBIT)
Annual Review of Microbiology 53129-54 (1999)
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Nature Reviews Genetics 7929-939 (2006)
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Generation of Mutant Libraries
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Labeled RNA Synthesis
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Transposon-Site Hybridization
Rich medium
Minimal medium
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Minimal medium
Rich medium
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Auxotrophic Mutations Identified by TraSH
Rich medium
Minimal medium
Ratio (Rich/Minimal)
Gene
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Superpathway of Leu, Val, and Ile biosynthesis
ilvA
leuD
leuB
BioCyc
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Superpathway of sulfate assimilation and Cys
biosynthesis
cysD
cysH
BioCyc
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EMBO Journal 155470-5479 (1996)
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Codon Usage of M. bovis

• TAT/TAC Tyr 2.06 (100 of Tyr)
• TTA/CTA Leu 0.63 (6.4 of Leu)
• ATA Ile 0.22 (5.2 of Ile)
• GTA Val 0.48 (5.6 of Val)
• TAA/TAG Stop 0.14 (46.6 of Stop)
• TATAAT -10 element

CBS
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Thanks for your attention!