RNA isolation

Total RNA may be isolated using TRIzol (from Thermo Fisher Scientific). Thousands of studies have been published using TRIzol. Alternatively, many dedicated RNA isolation kits are available. Suggestions include RNAeasy from Qiagen, miRCURY from Exiqon as well as a range of options from Promega and Thermo Fisher Scientific.

Tips

For final RNA elution or re-suspension, use water if possible. Water ensures no interference in any downstream RNA analyses. RNA stored in water is stable when frozen at -20°C to -80°C.

A portion of the total RNA sample (e.g. 1ug) can be run on a 1% agarose gel to ensure a non-degraded sample. Only the ribosomal RNA will be easily visible. A non-degraded sample is expected to display two distinct bands, a 28S ribosomal RNA band and an 18S ribosomal RNA band. The larger 26S band should be thicker.

Concentration analysis can be quickly performed using a Nanodrop (from Thermo Fisher Scientific). A Nanodrop takes up very little bench space, requires little training and maintenance and requires no reagents. A 260/280nm ratio of 2 suggests a reasonably pure RNA sample.

For links to molarity and dilution calculators see online science tools