They have proved satisfactory for isolating undegraded RNA that has been used to construct cDNA libraries, carry out in vitro translation and analyse RNA transcripts by Northern blotting. With all procedures for the isolation of intact RNA it is extremely important to ensure that equipment and reagents are RNAase free. Molecular biological grade reagents are used when available or alternatives of the highest quality. Due to the caustic nature of phenolic compounds care should be taken in the use of this reagent and extractions should be in a fume cupboard. Grind 1 g fresh weight of leaf tissue to a fine powder in liquid nitrogen using a mortar and pestle.
Re-extract the lower phase with an equal volume of extraction buffer and pool the aqueous phases.
Discard the supernatant. Any undissolved material will go into solution on addition of more water. However, it is important not to dilute the nucleic acid concentration substantially as the efficiency of subsequent LiCl precipitations can be reduced. RNA can now be selectively isolated using either LiCl or CsCl. After centrifugation, discard the supernatant.