Global Run-On Sequencing (GRO-seq) Library Preparation from Drosophila Ovaries

Rozhkov, N. V. (September 2015) Global Run-On Sequencing (GRO-seq) Library Preparation from Drosophila Ovaries. Methods Mol Biol, 1328. pp. 217-30. ISSN 1940-6029 (Electronic)1064-3745 (Linking)

URL: http://www.ncbi.nlm.nih.gov/pubmed/26324441
DOI: 10.1007/978-1-4939-2851-4_16

Abstract

In the past decade, deep-sequencing approaches have greatly improved our knowledge of the genome's potential and have become a crucial milestone for new discoveries in genomics. Transcription is the first step of gene expression; therefore, the detection and measurement of transcription rates is of great interest. Here, a detailed protocol for global run-on sequencing (GRO-seq) library preparation from Drosophila ovaries is described. The method relies on rapid isolation of nuclei with halted transcription, then restarting transcription in physiological conditions in the presence of a labeled nucleotide. The newly transcribed nascent RNA is then isolated and cloned using a small RNA cloning protocol. Although it is time-consuming, the global run-on method allows the user to profile the position, orientation and amount of transcriptionally engaged RNA polymerases across the genome, therefore providing a snapshot of genome-wide transcription.

Item Type: Paper
Subjects: organism description > animal > insect > Drosophila
organs, tissues, organelles, cell types and functions > cell types and functions > cell types > germ cell
organs, tissues, organelles, cell types and functions > cell types and functions > cell types > germ cell
organs, tissues, organelles, cell types and functions > cell types and functions > cell types > germ cell

Investigative techniques and equipment > assays > global run-on sequencing
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > transcriptomes
CSHL Authors:
Communities: CSHL labs > Hammell M. lab
Depositing User: Matt Covey
Date: 2 September 2015
Date Deposited: 04 Sep 2015 18:36
Last Modified: 04 Sep 2015 18:36
Related URLs:
URI: http://repository.cshl.edu/id/eprint/31735

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