Comprehensive identification of conditionally essential genes in mycobacteria

By Christopher M. Sassetti, D. H. Boyd, Eric J. Rubin. Proceedings of the National Academy of Sciences of the United States of America, 2001

622 citations recorded by the source corpus.

An increasing number of microbial genomes have been completely sequenced, and the identified genes are categorized based on their homology to genes of known function. However, the function of a large number of genes cannot be determined on this basis alone. Here, we describe a technique, transposon site hybridization (TraSH), which allows rapid functional characterization by identifying the complete set of genes required for growth under different conditions. TraSH combines high-density insertional mutagenesis with microarray mapping of pools of mutants. We have made large pools of independent transposon mutants in mycobacteria by using a mariner-based transposon and efficient phage transduction. By using TraSH, we have defined the set of genes required for growth of Mycobacterium bovis bacillus Calmette–Guérin on minimal but not rich medium. Genes of both known and unknown functions were identified. Of the genes with known functions, nearly all were involved in amino acid biosynthesis. TraSH is a powerful method for categorizing gene function that should be applicable to a variety of microorganisms.

Subfields: Ecology, Epidemiology, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/4facb512b851fb5a5c089fb31287bab4cbaa08ed

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Citations in the corpus

Genes required for mycobacterial growth defined by high density mutagenesis (2003, methodology): “In addition, many amino acid biosynthetic genes are required by mycobacteria regardless of culture condition (Sassetti et al., 2001), as these bacteria lack the ability to efficiently transport some exogenous compounds from the medium (Pavelka and Jacobs, 1999).”
Experimental Determination and System Level Analysis of Essential Genes in Escherichia coli MG1655 (2003, background): “terium tuberculosis (31), Haemophilus influenzae (1), and S.”
High-Resolution Phenotypic Profiling Defines Genes Essential for Mycobacterial Growth and Cholesterol Catabolism (2011, methodology): “Transposon mapping and identification of genes required for Mtb growth in vitro A library of transposon mutants consisting of approximately 10(5) independent insertion events was generated in the H37Rv strain of Mtb using a modified himar1 based transposon [5].”
Mycobacterium tuberculosis Pathogenesis and Molecular Determinants of Virulence (2003, background): “Ideally, the mariner transposon system (253) can be modified to incorporate signature tagging so that many potential mutants can be tested in vivo at the same time.”
Simultaneous assay of every Salmonella Typhi gene using one million transposon mutants. (2009, methodology): “1995) and transposon-site hybridization (TraSH) (Sassetti et al. 2001) are methods that make use of hybridization to identify genes disrupted by transposon insertions.”