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Identification and Analysis of Essential Genes in Streptococcus mutans with Transposon Sequencing

Part of the Methods in Molecular Biology book series (MIMB,volume 2377)

Abstract

Transposon sequencing (Tn-seq) has greatly accelerated the rate at which gene function can be profiled in microbial organisms. This technique has been applied to the study of the dental caries pathogen Streptococcus mutans where it has been used to generate large transposon mutant libraries. Coupled with high-throughput sequencing and bioinformatics tools, culture of these transposon mutant libraries has facilitated the identification of essential and conditional essential genes. In this chapter, we describe a procedure for performing Tn-seq studies in S. mutans that covers pooled transposon mutant construction, in vitro culture, and DNA library sequencing and data analysis.

Key words

  • Transposon mutagenesis
  • Essential genes
  • Streptococcus mutans
  • Tn-seq
  • Himar1
  • In vitro transposition

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Acknowledgements

We thank Nick Jakubovics at Newcastle University for help with the genomic DNA isolation protocol; Andrew Camilli at Tufts University for providing us the Tn-seq plasmids (pMalC9 and pMagellan6); Lin Zeng at the University of Florida for assistance with adapting the original Tn-seq protocol to work with S. mutans and help with editing this chapter; and David Moraga at the Interdisciplinary Center for Biotechnology Research at the University of Florida for help with troubleshooting and optimization of Tn-seq DNA sequencing.

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Correspondence to Robert C. Shields .

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Walker, A.R., Shields, R.C. (2022). Identification and Analysis of Essential Genes in Streptococcus mutans with Transposon Sequencing. In: Zhang, R. (eds) Essential Genes and Genomes. Methods in Molecular Biology, vol 2377. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1720-5_13

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  • DOI: https://doi.org/10.1007/978-1-0716-1720-5_13

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