Quantifying the ratio of spliceosome components assembled on pre-mRNA

Noa Neufeld, Yehuda Brody, Yaron Shav-Tal

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

3 Scopus citations

Abstract

RNA processing by the splicing machinery removes intronic sequences from pre-mRNA to generate mature mRNA transcripts. Many splicing events occur co-transcriptionally when the pre-mRNA is still associated with the transcription machinery. This mechanism raises questions regarding the number of spliceosomes associated with the pre-mRNA at a given time. In this protocol, we present a quantitative FISH approach that measures the ratio of intensities between two different spliceosomal components associated on a nascent mRNA, and compares to the number of introns in the mRNA, thereby calculating the number of spliceosome complexes assembled with each transcript.

Original languageEnglish
Title of host publicationSpliceosomal Pre-mRNA Splicing
Subtitle of host publicationMethods and Protocols
PublisherHumana Press Inc.
Pages257-269
Number of pages13
ISBN (Print)9781627039796
DOIs
StatePublished - 2014

Publication series

NameMethods in Molecular Biology
Volume1126
ISSN (Print)1064-3745

Funding

FundersFunder number
European Research Council
Seventh Framework Programme260441

    Keywords

    • Immunofluorescence
    • Intron
    • RNA FISH
    • Spliceosome

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