Super-resolution measurement of distance between transcription sites using RNA FISH with intronic probes
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Authors
Larkin, Joshua D.
Cook, Peter R.
Issue Date
2016
Type
Article
Language
Keywords
Colocalization , Diffraction limit , Inner-nuclear distance , Long gene , Nascent intronic RNA FISH , Super-resolution localization , Transcription location
Alternative Title
Abstract
Nascent transcripts being copied from specific human genes can be detected using RNA FISH (fluorescence in situ hybridization) with intronic probes, and the distance between two different nascent transcripts is often measured when studying structure-function relationships. Such distance measurements are limited by the resolution of the light microscope. Here we describe methods for measuring these distances in cultured cells with a precision of a few tens of nanometers, using equipment found in most laboratories (i.e., a wide-field fluorescence microscope equipped with a charged-coupled-device camera). Using images of pairs of transcripts that are often co-transcribed, we discuss how selection of cell type, design of FISH probes, image acquisition, and image processing affect the precision that can be achieved. (C) 2015 The Authors. Published by Elsevier Inc.
Description
Citation
Larkin, J. D., & Cook, P. R. (2016). Super-resolution measurement of distance between transcription sites using RNA FISH with intronic probes. Methods, 98, 150�"157. doi:10.1016/j.ymeth.2015.11.009
Publisher
License
Creative Commons Attribution 4.0 International
Journal
Volume
Issue
PubMed ID
ISSN
1046-2023