Author(s): Prudêncio, Pedro ; Rebelo, Kenny ; Grosso, Ana Rita ; Martinho, Rui Gonçalo ; Carmo-Fonseca, Maria
Date: 2020
Persistent ID: http://hdl.handle.net/10362/108217
Origin: Repositório Institucional da UNL
Project/scholarship: info:eu-repo/grantAgreement/FCT/5876/UID%2FBIM%2F04773%2F2013/PT;
Subject(s): Co-transcriptional splicing; mNET-seq; Nascent RNA; RNA polymerase II; Molecular Biology; Biochemistry, Genetics and Molecular Biology(all)
Fundação para a Ciência e Tecnologia (FCT)/ Ministério da Ciência, Tecnologia e Ensino Superior (MCTES) through Fundos do Orçamento de Estado (UID/BIM/50005/2019), and FCT/FEDER/POR Lisboa 2020, Programa Operacional Regional de Lisboa, PORTUGAL 2020 (LISBOA-01-0145-FEDER-016394).. R.G.M. is supported by FCT grants PTDC/BEX-BID/0395/2014 , PTDC/BIA-BID/28441/2017
Mammalian Native Elongating Transcript sequencing (mNET-seq) is a recently developed technique that generates genome-wide profiles of nascent transcripts associated with RNA polymerase II (Pol II) elongation complexes. The ternary transcription complexes formed by Pol II, DNA template and nascent RNA are first isolated, without crosslinking, by immunoprecipitation with antibodies that specifically recognize the different phosphorylation states of the polymerase large subunit C-terminal domain (CTD). The coordinate of the 3′ end of the RNA in the complexes is then identified by high-throughput sequencing. The main advantage of mNET-seq is that it provides global, bidirectional maps of Pol II CTD phosphorylation-specific nascent transcripts and coupled RNA processing at single nucleotide resolution. Here we describe the general pipeline to prepare and analyse high-throughput data from mNET-seq experiments.
