Comprehensive Essentiality Analysis of the Mycobacterium tuberculosis Genome via Saturating Transposon Mutagenesis

View/ Open
Author
DeJesus, Michael A.
Xu, Weizhen
Park, Sae Woong
Long, Jarukit E.
Schnappinger, Dirk
Ehrt, Sabine
Sassetti, Christopher M.
Ioerger, Thomas R.
Note: Order does not necessarily reflect citation order of authors.
Published Version
https://doi.org/10.1128/mBio.02133-16Metadata
Show full item recordCitation
DeJesus, M. A., E. R. Gerrick, W. Xu, S. W. Park, J. E. Long, C. C. Boutte, E. J. Rubin, et al. 2017. “Comprehensive Essentiality Analysis of the Mycobacterium tuberculosis Genome via Saturating Transposon Mutagenesis.” mBio 8 (1): e02133-16. doi:10.1128/mBio.02133-16. http://dx.doi.org/10.1128/mBio.02133-16.Abstract
ABSTRACT For decades, identifying the regions of a bacterial chromosome that are necessary for viability has relied on mapping integration sites in libraries of random transposon mutants to find loci that are unable to sustain insertion. To date, these studies have analyzed subsaturated libraries, necessitating the application of statistical methods to estimate the likelihood that a gap in transposon coverage is the result of biological selection and not the stochasticity of insertion. As a result, the essentiality of many genomic features, particularly small ones, could not be reliably assessed. We sought to overcome this limitation by creating a completely saturated transposon library in Mycobacterium tuberculosis. In assessing the composition of this highly saturated library by deep sequencing, we discovered that a previously unknown sequence bias of the Himar1 element rendered approximately 9% of potential TA dinucleotide insertion sites less permissible for insertion. We used a hidden Markov model of essentiality that accounted for this unanticipated bias, allowing us to confidently evaluate the essentiality of features that contained as few as 2 TA sites, including open reading frames (ORF), experimentally identified noncoding RNAs, methylation sites, and promoters. In addition, several essential regions that did not correspond to known features were identified, suggesting uncharacterized functions that are necessary for growth. This work provides an authoritative catalog of essential regions of the M. tuberculosis genome and a statistical framework for applying saturating mutagenesis to other bacteria.Other Sources
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5241402/pdf/Terms of Use
This article is made available under the terms and conditions applicable to Other Posted Material, as set forth at http://nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of-use#LAACitable link to this page
http://nrs.harvard.edu/urn-3:HUL.InstRepos:30371135
Collections
- FAS Scholarly Articles [17837]
- HMS Scholarly Articles [18278]
- SPH Scholarly Articles [6399]
Contact administrator regarding this item (to report mistakes or request changes)