title: Mapping 3' mRNA isoforms on a genomic scale.
abstract: Most eukaryotic genes are transcribed into mRNAs with alternative poly(A) sites. Emerging evidence suggests that mRNA isoforms with alternative poly(A) sites can perform critical regulatory functions in numerous biological processes. In recent years, a number of strategies utilizing high-throughput sequencing technologies have been developed to aid in the identification of genome-wide poly(A) sites. This unit describes a modified protocol for a recently published 3'READS (3' region extraction and deep sequencing) method that accurately identifies genome-wide poly(A) sites and that can be used to quantify the relative abundance of the resulting 3' mRNA isoforms. This approach minimizes nonspecific sequence reads due to internal priming and typically yields a high percentage of sequence reads that are ideally suited for accurate poly(A) identification.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Yi
authors.initials: Y
authors.lastname: Jin
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Joseph V
authors.initials: JV
authors.lastname: Geisberg
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Zarmik
authors.initials: Z
authors.lastname: Moqtaderi
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Zhe
authors.initials: Z
authors.lastname: Ji
authors.affiliation: Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers New Jersey Medical School, Newark, New Jersey.
authors.forename: Mainul
authors.initials: M
authors.lastname: Hoque
authors.affiliation: Department of Microbiology, Biochemistry and Molecular Genetics, Rutgers New Jersey Medical School, Newark, New Jersey.
authors.forename: Bin
authors.initials: B
authors.lastname: Tian
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts.
authors.forename: Kevin
authors.initials: K
authors.lastname: Struhl
mesh_terms: D020413:3' Untranslated Regions; D059014:High-Throughput Nucleotide Sequencing; D008967:Molecular Biology / Q000379:methods*; D011061:Poly A / Q000235:genetics*; D059368:RNA Isoforms / Q000032:analysis* / Q000235:genetics* / Q000302:isolation & purification
chemical_list: D020413:3' Untranslated Regions; D059368:RNA Isoforms; D011061:Poly A
keywords: 3′ mRNA isoforms; RNA sequencing; poly(A) sites; polyadenylation
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