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Saccharomyces cerevisiae 3' poly(A) site mapping

ABSTRACT: The use of alternative polyadenylation sites is common and affects the post-transcriptional fate of mRNA, including its stability, localization, and translation. Here we present a method for genome-wide and strand-specific mapping of poly(A) sites and quantification of RNA levels at unprecedented efficiency by using an on-cluster dark T-fill procedure on the Illumina sequencing platform. Our method outperforms former protocols in quality and throughput, and reveals new insights into polyadenylation in Saccharomyces cerevisiae. Experimental benchmark of five different protocols (3tfill, bpmI, internal, rnaseq and yoon) for genome-wide identification of polyadenylation sites in Saccharomyces cerevisiae and transcript quantification. RNA was extracted from WT cells grown in glucose (ypd) or galactose (ypgal) as carbon source. The same RNA was used for 3 independent library constructions (technical replicates, rep).

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Aino Järvelin

PROVIDER: E-GEOD-40110 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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An efficient method for genome-wide polyadenylation site mapping and RNA quantification.

Wilkening Stefan S Pelechano Vicent V Järvelin Aino I AI Tekkedil Manu M MM Anders Simon S Benes Vladimir V Steinmetz Lars M LM

Nucleic acids research 20130107 5

The use of alternative poly(A) sites is common and affects the post-transcriptional fate of mRNA, including its stability, subcellular localization and translation. Here, we present a method to identify poly(A) sites in a genome-wide and strand-specific manner. This method, termed 3'T-fill, initially fills in the poly(A) stretch with unlabeled dTTPs, allowing sequencing to start directly after the poly(A) tail into the 3'-untranslated regions (UTR). Our comparative analysis demonstrates that it ...[more]

PMID: 23295673

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