Detection of Nascent RNA Transcripts by Fluorescence In Situ Hybridization
The development of cellular diversity within any organism depends on the timely and correct expression of differing subsets of genes within each tissue type. Many techniques exist which allow a global, average analysis of RNA expression; however, RNA-FISH permits the sensitive detection of specific transcripts within individual cells while preserving the cellular morphology. The technique can provide insight into the spatial and temporal organization of gene transcription as well the relationship of gene expression and mature RNA distribution to nuclear and cellular compartments. It can also reveal the intercellular variation of gene expression within a given tissue. Here, we describe RNA-FISH methodologies that allow the detection of nascent transcripts within the cell nucleus as well as protocols that allow the detection of RNA alongside DNA or proteins. Such techniques allow the placing of gene transcription within a functional context of the whole cell.
Key wordsTranscription RNA-FISH RNA-immunoFISH RNA-DNA-FISH RNA Methods Nascent transcript RNA probes Gene expression
We thank the Grosveld Lab in Rotterdam, particularly Peter Fraser, Joost Gribnau, Tolleiv Trimborn, and Mark Wijgerde for initial support with setting up RNA-FISH as a technique in our laboratory. We thank Jackie Sloane-Stanley and Sue Butler for provision of cells. Our work is funded by the Medical Research Council.
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