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

  • Stéphane Poulain
  • Sachi Kato
  • Ophélie Arnaud
  • Jean-Étienne Morlighem
  • Makoto Suzuki
  • Charles Plessy
  • Matthias Harbers
Part of the Methods in Molecular Biology book series (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.

Key words

Cap analysis gene expression CAGE nanoCAGE CAGEscan RNA Transcription Start Sites TSS Expression profiling Template switching Tagmentation Multiplexing Unique molecular identifiers UMI 

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Stéphane Poulain
    • 1
  • Sachi Kato
    • 1
    • 2
  • Ophélie Arnaud
    • 1
  • Jean-Étienne Morlighem
    • 2
    • 3
  • Makoto Suzuki
    • 2
    • 4
  • Charles Plessy
    • 1
    • 2
  • Matthias Harbers
    • 1
    • 2
  1. 1.Division of Genomic TechnologiesRIKEN Center for Life Science TechnologiesYokohamaJapan
  2. 2.RIKEN Omics Science Center (OSC)YokohamaJapan
  3. 3.Laboratory of Biochemistry and Biotechnology, Institute for Marine SciencesFederal University of CearaFortalezaBrazil
  4. 4.DNAFORM, Inc.YokohamaJapan

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