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mRNA-seq with agnostic splice site discovery for nervous system transcriptomics tested in chronic pain

  • mRNA-seq is a paradigm-shifting technology because of its superior sensitivity and dynamic range and its potential to capture transcriptomes in an agnostic fashion, i.e., independently of existing genome annotations. Implementation of the agnostic approach, however, has not yet been fully achieved. In particular, agnostic mapping of pre-mRNA splice sites has not been demonstrated. The present study pursued dual goals: (1) to advance mRNA-seq bioinformatics toward unbiased transcriptome capture and (2) to demonstrate its potential for discovery in neuroscience by applying the approach to an in vivo model of neurological disease. We have performed mRNA-seq on the L4 dorsal root ganglion (DRG) of rats with chronic neuropathic pain induced by spinal nerve ligation (SNL) of the neighboring (L5) spinal nerve. We found that 12.4% of known genes were induced and 7% were suppressed in the dysfunctional (but anatomically intact) L4 DRG 2 wk after SNL. These alterations persisted chronically (2 mo). Using a read cluster classifier with strong test characteristics (ROC area 97%), we discovered 10,464 novel exons. A new algorithm for agnostic mapping of pre-mRNA splice junctions (SJs) achieved a precision of 97%. Integration of information from all mRNA-seq read classes including SJs led to genome reannotations specifically relevant for the species used (rat), the anatomical site studied (DRG), and the neurological disease considered (pain); for example, a 64-exon coreceptor for the nociceptive transmitter substance P was identified, and 21.9% of newly discovered exons were shown to be dysregulated. Thus, mRNA-seq with agnostic analysis methods appears to provide a highly productive approach for in vivo transcriptomics in the nervous system.

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Metadaten
Author:Paul Hammer, Michaela S. Banck, Ronny Amberg, Cheng Wang, Gabriele Petznick, Shujun Luo, Irina Khrebtukova, Gary P. Schroth, Peter Beyerlein, Andreas S. Beutler
URN:urn:nbn:de:kobv:526-opus4-6446
DOI:https://doi.org/10.1101/gr.101204.109
ISSN:1549-5469
Parent Title (English):Genome Research
Document Type:Article
Language:English
Year of Publication:2010
Publishing Institution:Technische Hochschule Wildau
Release Date:2016/07/06
Volume:20
Issue:6
Page Number:15
Source:Paul Hammer, Michaela S. Banck, Ronny Amberg, et al. mRNA-seq with agnostic splice site discovery for nervous system transcriptomics tested in chronic pain. Genome Res. 2010 20: 847-860 originally published online May 7, 2010. Access the most recent version at doi: 10.1101/gr.101204.109
Faculties an central facilities:Fachbereich Ingenieurwesen / Wirtschaftsingenieurwesen (bis 8/2014)
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Licence (German):Creative Commons - CC BY-NC 3.0 - Namensnennung-Nicht kommerziell 3.0 Unported
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