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Promoter Associated RNA pp 57–109Cite as

NanoCAGE: A Method for the Analysis of Coding and Noncoding 5′-Capped Transcriptomes

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1543))

Abstract

Transcripts in all eukaryotes are characterized by the 5′-end specific cap structure in mRNAs. Cap Analysis Gene Expression or CAGE makes use of these caps to specifically obtain cDNA fragments from the 5′-end of RNA and sequences those at high throughput for transcript identification and genome-wide mapping of transcription start sites for coding and noncoding genes. Here, we provide an improved version of our nanoCAGE protocol that has been developed for preparing CAGE libraries from as little as 50 ng of total RNA within three standard working days. Key steps in library preparation have been improved over our previously published protocol to obtain libraries having a good 5′-end selection and a more equal size distribution for higher sequencing efficiency on Illumina MiSeq and HiSeq sequencers. We recommend nanoCAGE as the method of choice for transcriptome profiling projects even from limited amounts of RNA, and as the best approach for genome-wide mapping of transcription start sites within promoter regions.

The original version of this chapter was revised. The erratum to this chapter is available at: DOI 10.1007/978-1-4939-6716-2_17

An erratum to this chapter can be found at http://dx.doi.org/10.1007/978-1-4939-6716-2_17

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Acknowledgments

We thank Alexandre Fort for critically reading the manuscript and his helpful comments, and Laia Masvidal Sanz for helpful discussions and suggestions on the nanoCAGE protocol. This work was founded by a Research Grant from the Japanese Ministry of Education, Culture, Sports, Science and Technology (MEXT) to the RIKEN Center for Life Science Technologies, and a JSPS Grant-in-Aid for Young Scientists A (number 25710018).

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Authors and Affiliations

  1. Division of Genomic Technologies, RIKEN Center for Life Science Technologies, 1-7-22, Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa, 230-0045, Japan

    Stéphane Poulain, Sachi Kato, Ophélie Arnaud, Charles Plessy & Matthias Harbers

  2. RIKEN Omics Science Center (OSC), 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, 230-0045, Japan

    Sachi Kato, Jean-Étienne Morlighem, Makoto Suzuki, Charles Plessy & Matthias Harbers

  3. Laboratory of Biochemistry and Biotechnology, Institute for Marine Sciences, Federal University of Ceara, Av. da Abolição, 3207—Meireles, Fortaleza, CE, 60165-081, Brazil

    Jean-Étienne Morlighem

  4. DNAFORM, Inc., Leading Venture Plaza 2, 75-1 Ono-cho, Tsurumi-ku, Yokohama, Kanagawa, 230-0046, Japan

    Makoto Suzuki

Authors
  1. Stéphane Poulain
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  2. Sachi Kato
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  3. Ophélie Arnaud
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  4. Jean-Étienne Morlighem
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  5. Makoto Suzuki
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  6. Charles Plessy
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  7. Matthias Harbers
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Corresponding authors

Correspondence to Charles Plessy or Matthias Harbers .

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Editors and Affiliations

  1. Experimental Therapeutics Group, Institute of Oncology Research, Bellinzona, Switzerland

    Sara Napoli

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Poulain, S. et al. (2017). NanoCAGE: A Method for the Analysis of Coding and Noncoding 5′-Capped Transcriptomes. In: Napoli, S. (eds) Promoter Associated RNA. Methods in Molecular Biology, vol 1543. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6716-2_4

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  • DOI: https://doi.org/10.1007/978-1-4939-6716-2_4

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6714-8

  • Online ISBN: 978-1-4939-6716-2

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