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RNA Preparation and RNA-Seq Bioinformatics for Comparative Transcriptomics

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Microbial Steroids

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2704))

Abstract

The principal transcriptome analysis is the determination of differentially expressed genes across experimental conditions. For this, the next-generation sequencing of RNA (RNA-seq) has several advantages over other techniques, such as the capability of detecting all the transcripts in one assay over RT-qPCR, such as its higher accuracy and broader dynamic range over microarrays or the ability to detect novel transcripts, including non-coding RNA molecules, at nucleotide-level resolution over both techniques. Despite these advantages, many microbiology laboratories have not yet applied RNA-seq analyses to their investigations. The high cost of the equipment for next-generation sequencing is no longer an issue since this intermediate part of the analysis can be provided by commercial or central services. Here, we detail a protocol for the first part of the analysis, the RNA extraction and an introductory protocol to the bioinformatics analysis of the sequencing data to generate the differential expression results.

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Acknowledgments

This work was supported by a grant of the European Union program ERA-IB [MySterI (EIB.12.010)] through the APCIN call of the Spanish Ministry of Economy and Competitiveness (MINECO, Spain) (PCIN-2013-024-C02-01). The authors thank the European Union program ERA-IB; the Spanish Ministry of Economy and Competitiveness (MINECO, Spain); the MySter I Consortium (INBIOTEC, Pharmins Ltd., University of York, SINTEF, Technische Universität Dortmund, and Gadea Biopharma S.L.), and the Syntheroids project (ERA-CoBioTech 1st call; founded through the APCIN call of the Spanish Ministry of Science, Innovation and Universities; ID: PCI2018-093066). They also thank J. Merino, B. Martín and A. Casenave for their excellent technical assistance and the degree students of the group A. Morales and P. González.

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Authors and Affiliations

  1. Área de Microbiología, Departamento de Biología Molecular, Facultad de Ciencias Biológicas y Ambientales, Universidad de León, León, Spain

    Antonio Rodríguez-García

  2. Instituto de Biotecnología de León, INBIOTEC, León, Spain

    Antonio Rodríguez-García, Alberto Sola-Landa & Carlos Barreiro

  3. Fundación Cesefor, León, Spain

    Alberto Sola-Landa

  4. Área de Bioquímica y Biología Molecular, Departamento de Biología Molecular, Facultad de Veterinaria, Universidad de León, León, Spain

    Carlos Barreiro

Authors
  1. Antonio Rodríguez-García
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  2. Alberto Sola-Landa
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  3. Carlos Barreiro
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Corresponding author

Correspondence to Antonio Rodríguez-García .

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Editors and Affiliations

  1. Área de Bioquímica y Biología Molecular, Departamento de Biología Molecular, Facultad de Veterinaria, Universidad de León, León, Spain

    Carlos Barreiro

  2. Department of Biotechnology, Curia Spain, León, Spain

    José-Luis Barredo

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Rodríguez-García, A., Sola-Landa, A., Barreiro, C. (2023). RNA Preparation and RNA-Seq Bioinformatics for Comparative Transcriptomics. In: Barreiro, C., Barredo, JL. (eds) Microbial Steroids. Methods in Molecular Biology, vol 2704. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3385-4_6

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  • DOI: https://doi.org/10.1007/978-1-0716-3385-4_6

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3384-7

  • Online ISBN: 978-1-0716-3385-4

  • eBook Packages: Springer Protocols

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