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RiboTag : Ribosomal tagging strategy to analyze cell type specific mRNA expression in vivo
Sanz Iglesias, Elisenda (Universitat Autònoma de Barcelona. Institut de Neurociències)
Bean, Jonathan C. (University of Washington. Department of Pharmacology)
Carey, Daniel P. (University of Washington. Department of Pharmacology)
Quintana Romero, Albert (Universitat Autònoma de Barcelona. Institut de Neurociències)
McKnight, G. Stanley (University of Washington. Department of Pharmacology)

Date: 2019
Abstract: Ribosome tagging has become a very useful in vivo approach for analyzing gene expression and mRNA translation in specific cell types that are difficult and time consuming to isolate by conventional methods. The approach is based on selectively expressing an HA-tagged ribosomal protein in a target cell type and then using antibodies against HA to purify the polysomes and associated mRNAs from the target cell. The original approach makes use of a mouse line (RiboTag) harboring a modified allele of Rpl22 (Rpl22-HA) that is induced by the action of Cre recombinase. The Rpl22-HA gene can also be introduced into the animal by stereotaxic injections of an AAV-DIO-Rpl22-HA that is then activated in Cre-expressing cells. Both methods for tagging ribosomes facilitate the immunoprecipitation of ribosome bound mRNAs and their analysis by qRT-PCR or RNA-Seq. This protocol will discuss the technical procedures and describe important considerations relevant to the analysis of the data.
Note: Número d'acord de subvenció EC/H2020/658352
Note: Número d'acord de subvenció EC/H2020/638106
Note: Número d'acord de subvenció MINECO/RyC-2012-11873
Note: Número d'acord de subvenció MINECO/SAF2014-57981P
Note: Número d'acord de subvenció MINECO/SAF2017-88108-R
Note: Número d'acord de subvenció AGAUR/2017/SGR-323
Rights: Tots els drets reservats
Language: Anglès.
Document: article ; recerca ; submittedVersion
Subject: Gene expression ; Ribosome tagging ; Neuronal transcripts ; Polyribosome ; Immunoprecipitation
Published in: Current Protocols in Neuroscience, 2019 , ISSN 1934-8576

DOI: 10.1002/cpns.77


Preprint
58 p, 970.8 KB

The record appears in these collections:
Research literature > UAB research groups literature > Research Centres and Groups (scientific output) > Health sciences and biosciences > Institut de Neurociències (INc)
Articles > Research articles
Articles > Published articles

 Record created 2019-05-29, last modified 2019-07-15



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