UNIT 5.13 Fluorescent In Situ Transcription in Cells and Tissues
Published Online: 1 MAY 2001
Copyright © 2003 by John Wiley and Sons, Inc.
Lab Protocol Title
Current Protocols in Neuroscience
How to Cite
Ariano, M. A. 2001. Fluorescent In Situ Transcription in Cells and Tissues. Current Protocols in Neuroscience. 8:5.13:5.13.1–5.13.10.
- Published Online: 1 MAY 2001
- Published Print: AUG 1999
This method assesses cellular mRNA transcripts in tissue sections and cell cultures using unique short anti-sense primers directed against sequences in particular protein(s). The unlabeled synthetic cDNA oligonucleotide primers are extended complementary to a sense mRNA transcript using reverse transcriptase and labeled through incorporation of a fluorescent-labeled dUTP nucleotide base. The new cDNA will be synthesized upstream from the point of primer hybridization, and has a specific activity of fluorescent labeling dependent upon the length of the template mRNA from the primer location to the 5'-terminus. This procedure provides rapid detection of low abundance mRNA messages that can be related to other cellular protein components, labeled experimentally with alternative fluorochromes.