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)
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: | https://repository.cshl.edu/id/eprint/31735 |
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