Protocol

Circadian Rhythms

Volume 362 of the series Methods in Molecular Biology™ pp 315-327

RNA Extraction From Mammalian Tissues

  • Stuart N. PeirsonAffiliated withDivision of Circadian and Visual Neuroscience, University of Oxford
  • , Jason N. ButlerAffiliated withDivision of Circadian and Visual Neuroscience, University of Oxford

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Abstract

Purification of intact RNA is the primary step of many molecular biology techniques, including Northern blotting, RNase protection, quantitative polymerase chain reaction, and microarray assays. RNA extraction is typically conducted using either a phenol-choloroform or a solid phase method. This article concentrates primarily on the former approach, which is highly versatile, and is easily adapted to different tissues ranging from whole organs down to submillimeter biopsy punches.

The major problem with RNA extraction is the ubiquitous nature of RNases, enzymes that rapidly degrade RNA. RNases may be difficult to eliminate, although with care and appropriate countermeasures, reproducible extraction of high-quality RNA from important biological samples should be attainable.

This article focuses on the isolation of RNA from the tissue collection step, homogenization all the way through to the quantification of the purified nucleic acid, providing guidelines for the prevention of the problems associated with RNAse contamination.

Key Words

RNases quantification homogenization microfluidics