Abstract
During the construction of random transposon mutagenesis libraries, four essential statistical issues arise: (1) Computing basic probability results for number of open reading frame knockouts. (2) Estimating the number of new open reading frames that will be knockouts in the next set of clones. (3) Estimating the number of essential open reading frames. (4) Computing the probability that an open reading frame is essential given the distribution of insertions. This chapter examines these issues and evaluates potential solutions using three different approaches: Efron and Thisted’s estimator, Will and Jacobs’s parametric bootstrap, and Blades and Broman’s Gibbs sampler. In doing so, this chapter provides guidance for using the R statistical project to solve these problems.
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References
Jacobs, M. A., Alwood, A., Thaipisuttikul, I., Spencer, D., Haugen, E., Ernst, S., et al. (2003) Comprehensive transposon mutant library of Pseudomonas aeruginosa. Proc. Natl. Acad. Sci. U.S.A. 100, 14339–14344.
Hayes, F. (2003) Transposon-based strategies for microbial functional genomics and proteomics. Annu. Rev. Genet. 37, 3–29.
Fraser, C. M. (2004) A genomics-based approach to biodefence preparedness. Nature Rev. Genet. 5, 23–33.
Gerdes, S. Y., Scholle, M. D., Campbell, J. W., Balázsi, G., Ravasz, E., Daugherty, M. D., et al. (2003) Experimental determination and system level analysis of essential genes in Escherichia coli MG1655. J. Bacteriol. 185, 5673–5684.
Giaever, G., Chu, A. M., Ni, L., Connelly, C., Riles, L., Véronneau, S., et al. (2002) Functional profiling of Saccharomyces cerevisiae genome. Nature 418, 387–391.
Hutchison IC. A. III, Peterson, S. N., Gill, S. R., Cline, R. T., White, O., Fraser, C. M., et al. (1999) Global transposon mutagenesis and a minimal mycoplasma genome. Science. 286, 2165–2169.
Kobayashi, K., Ehrlich, S. D., Albertini, A., Amati, G., Andersen, K. K., Arnaud, M., et al. Essential Bacillus subtilis genes. Proc. Natl. Acad. Sci. U.S.A. 100, 4678–4683.
Lamichhane, G., Zignol, M., Blades, N. J., Geiman, D. E., Dougherty, A., Grosset, J., et al. (2003) A postgenomic method for predicting essential genes at subsaturation levels of mutagenesis: application to Mycobacterium tuberculosis. Proc. Natl. Acad. Sci. U.S.A. 100, 7213–7218.
Efron, B., and Thisted, R. (1976) Estimating the number of unseen species: How many words did Shakespeare know? Biometrika. 63, 435–447.
Blades, N. J., and Broman, K. W. (2002) Estimating the number of essential genes in a genome by random tranposon mutagenesis. Technical Report MS02-20. Baltimore, MD: Johns Hopkins University.
Johnson, N. L., and Kotz, S. (1977) Urn Models and Their Application: An Approach to Modern Discrete Probability Theory. New York: John Wiley & Sons.
Davison, A. C., and Hinkley, D. V. (1997) Bootstrap Methods and their Applications. Cambridge: Cambridge University Press.
Broman, K. W. (2004) negenes: Estimating the Number of Essential Genes in a Genome. R package version 0.98–5. Available at http://www.biostat.jhsph.edu/~kbroman/software/negenes.html.
R Development Core Team (2005) R: A Language and Environment for Statistical Computing. Vienna, Austria: R Foundation for Statistical Computing. Available at http://www.R-project.org.
Krause, A., and Olson, M. (2000) The Basics of S-PLUS. New York: Springer-Verlag.
Will, O., and Jacobs, M. A. (2006) Estimating the number of essential genes in random transposon mutagenesis libraries. Available at http://arxiv.org/abs/q-bio.OT/0608005.
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Will, O. (2008). Statistical Methods for Building Random Transposon Mutagenesis Libraries. In: Osterman, A.L., Gerdes, S.Y. (eds) Microbial Gene Essentiality: Protocols and Bioinformatics. Methods in Molecular Biology™, vol 416. Humana Press. https://doi.org/10.1007/978-1-59745-321-9_22
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DOI: https://doi.org/10.1007/978-1-59745-321-9_22
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