RNA extraction tips

Last update: April 26, 2016

A vulnerable molecule

red drop in a tube with waterRNA extraction from biological samples is complicated by the vulnerability of the RNA molecule itself and by the ubiquitous presence of ribonuclease enzymes in cells, tissues, and environment which can rapidly degrade RNA (for instance, RNase 7, a member of the RNase A superfamily is secreted by human skin and serves as a potent antipathogen defence). With respect to DNA, the RNA molecule is very unstable because it is a single strand, whereas DNA is double stranded. In addition, certain RNases can be extremely hardy and inactivating them is difficult compared to neutralizing DNases.

Extraction methods

Several methods are used in molecular biology to isolate RNA. The most common is the guanidinium thiocyanate-phenol-chloroform method (the reagent is sold by Sigma-Aldrich as TRI Reagent, by Invitrogen as TRIzol, by Bioline as Trisure, and by Tel-Test as STAT-60).

RNA can be extracted from several different types of tissues: blood, FFPE, cultered cells, various tissues and saliva. For RNA extraction from saliva there are some commercial kits available that stabilize the RNA molecule immediately after sampling (e.g. RNeasy Protect Saliva Mini Kit – Qiagen: according to the manufacturer, it is possible with this kit to stabilize and ship the samples at 37°C for 1 day, 15–25°C for 14 days, or 2–8°C for 4 weeks before RNA purification).

Studying the transcriptome

However it is important to remember that the pool of RNA molecules (the so called transcriptome) is different from tissue to tissue. So, even when saliva sampling is available or preferrable, it may be unsuful simply because the genes of interest are not expressed in buccal cells.

References: https://en.wikipedia.org/wiki/RNA_extractionPMID: 23564756

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