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  • Analysis of messenger RNA expression by in situ hybridization using RNA probes synthesized via in vitro transcription.

Analysis of messenger RNA expression by in situ hybridization using RNA probes synthesized via in vitro transcription.

Methods (San Diego, Calif.) (2010-08-12)
Bradley S Carter, Jonathan S Fletcher, Robert C Thompson
ABSTRACT

The analysis of the spatial patterning of mRNA expression is critically important for assigning functional and physiological significance to a given gene product. Given the tens of thousands of mRNAs in the mammalian genome, a full assessment of individual gene functions would ideally be overlaid upon knowledge of the specific cell types expressing each mRNA. In situ hybridization approaches represent a molecular biological/histological method that can reveal cellular patterns of mRNA expression. Here, we present detailed procedures for the detection of specific mRNAs using radioactive RNA probes in tissue sections followed by autoradiographic detection. These methods allow for the specific and sensitive detection of spatial patterns of mRNA expression, thereby linking mRNA expression with cell type and function. Radioactive detection methods also facilitate semi-quantitative analyses of changes in mRNA gene expression.

MATERIALS
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Product Description

Sigma-Aldrich
Cresyl Violet acetate, certified by the Biological Stain Commission
Sigma-Aldrich
Dextran sulfate sodium salt from Leuconostoc spp., for molecular biology, average Mw >500,000 (dextran starting material), contains 0.5-2% phosphate buffer