Abstract
The adaptation of Legionella pneumophila to the different conditions it encounters in the environment and in the host is governed by a complex regulatory system. Current knowledge of these regulatory networks and the transcriptome responses of L. pneumophila is mainly based on microarray analysis and limited to transcriptional products of annotated protein-coding genes. The application of the Next-Generation Sequencing (NGS) technology allows now genome-wide strand-specific sequencing and accurate determination of all expressed regions of the genome to reveal the complete transcriptional network and the dynamic interplay of specific regulators on a genome-wide level. NGS-based techniques promote deeper understanding of the global transcriptional organization of L. pneumophila by identifying transcription start sites (TSS), alternative TSS and operon organization, noncoding RNAs, antisense RNAs, and 5′-/3′-untranslated regions. In this chapter we describe the construction of cDNA libraries for (1) RNA deep sequencing (RNA-seq) and (2) TSS mapping using the Illumina technology.
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Acknowledgments
This work received support from the Institut Pasteur, the Centre national de la recherché scientifique (CNRS), and the Institut Carnot-Pasteur MI, the Labex project “IBEID” and from the ANR-10-PATH-004 project, in the frame of ERA-Net PathoGenoMics.
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Sahr, T., Buchrieser, C. (2013). cDNA Library Construction for Next-Generation Sequencing to Determine the Transcriptional Landscape of Legionella pneumophila . In: Buchrieser, C., Hilbi, H. (eds) Legionella. Methods in Molecular Biology, vol 954. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-161-5_34
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DOI: https://doi.org/10.1007/978-1-62703-161-5_34
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Publisher Name: Humana Press, Totowa, NJ
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