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Targeted RNA Sequencing Assay to Characterize Gene Expression and Genomic Alterations
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TARDIS, a targeted RNA directional sequencing method for rare RNA discovery.

Maximiliano M Portal1, Valeria Pavet1, Cathie Erb1

  • 1Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Equipe Labellisée Ligue Contre le Cancer, Centre National de la Recherche Scientifique UMR 7104, Institut National de la Santé et de la Recherche Médicale U964, University of Strasbourg, Illkirch, France.

Nature Protocols
|October 30, 2015
PubMed
Summary

Targeted-RNA Directional Sequencing (TARDIS) enables the identification of rare transcripts. This method interrogates specific RNA subsets, overcoming limitations of standard RNA sequencing for novel transcript discovery.

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Area of Science:

  • Molecular Biology
  • Genomics
  • Transcriptomics

Background:

  • High-throughput sequencing reveals numerous novel RNAs.
  • Identifying rare transcripts is challenging due to technical limitations in RNA sequencing library preparation and platform performance.

Purpose of the Study:

  • To introduce Targeted-RNA Directional Sequencing (TARDIS), a novel hybridization-based method.
  • To enable independent interrogation of specific RNA subsets within the transcriptome, irrespective of transcript characteristics.

Main Methods:

  • TARDIS is a modular protocol involving DNA trap generation, RNA purification, DNA trap-based RNA capture, and library construction.
  • The method utilizes hybridization for targeted RNA capture.
  • Coupling RNA capture with strand-specific RNA sequencing.

Main Results:

  • TARDIS allows robust identification and reconstruction of novel transcripts.
  • It facilitates the definition of sense and antisense RNA pairs.
  • Concomitant analysis of long and small RNA pools enables inference of precursor-product relationships.

Conclusions:

  • TARDIS is an effective method for interrogating specific RNA subsets.
  • The protocol overcomes limitations of standard RNA sequencing for rare and novel transcript discovery.
  • TARDIS provides a comprehensive approach to transcriptomic analysis, including novel transcript identification and precursor-product relationship inference.