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  • Rapid, high-quality and epidermal-specific isolation of RNA from human skin.

Rapid, high-quality and epidermal-specific isolation of RNA from human skin.

Experimental dermatology (2007-02-09)
Andrea Trost, Johann W Bauer, Christoph Lanschützer, Martin Laimer, Michael Emberger, Helmut Hintner, Kamil Onder
ABSTRACT

As global transcriptome analyses with a growing demand on layer-specific applications are widely used in cutaneous biology, we investigated the effect of established and optimized dermo-epidermal separation methods on the quality of RNA. We compared enzymatic separation with dispase, chemical separation with 1 M sodium chloride and heat separation to a treatment with 3.8% ammonium thioyanate. The impact of freezing as well as the addition of 10 mM aurintricarboxylic acid was considered in the evaluation of the amount and quality of isolated RNA from dermis and epidermis. Using the low abundant gene kallikrein 12 for real-time PCR analysis, we were able to demonstrate the superior RNA quality after dermo-epidermal separation using 3.8% ammonium thiocyanate. In addition to the time effectiveness this separation technique promises dermal and epidermal purity and is therefore the method of choice for producing high-quality RNA for genome-wide dermal and epidermal transcriptional analysis.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Ammonium thiocyanate solution, 0.1 M, 1/10 N
Sigma-Aldrich
Ammonium thiocyanate, SAJ special grade, ≥99%
Sigma-Aldrich
Ammonium thiocyanate, powder
Sigma-Aldrich
Ammonium thiocyanate, ACS reagent, ≥97.5%
Supelco
Ammonium thiocyanate, 99.99% trace metals basis