All rights of reproduction in any form reserved. 1% 8-hydroxyquinoline (w/v), is a useful stock reagent. Wrapped in foil and stored at 4 °, this reagent can be kept for up to 1 month. If no 8-hydroxyquinoline is added, the eventual discoloring of the phenol indicates that it is extensively oxidized and should be discarded. If the commercially supplied phenol is either crystalline or has a yellow or pink coloration, it is necessary to purify it further by redistillation. The coloration is indicative of the presence of phenolic oxidation products (quinones, diacids, and others) which form during storage.

Chem. 252, 5904 (1977). 38 I S O L A T I N G A N D C H A R A C T E R I Z I N G N U C L E I C ACIDS [4] phase and interphase. Heparin, another common ribonuclease inhibitor (see this volume [24]), cannot be removed by extracting with phenol. Physical Considerations. Extractions of nucleic acids from biological sources are best carried out as quickly as possible in ice with refrigerated buffers to minimize nuclease activity. Only when SDS is present would a higher temperature (10°) be required to prevent precipitation of the detergent.

23-25 Its activity is critically dependent upon inclusion of a minimum of 1 mM dithiothreitol, and care must be taken not to expose the RNasin-RNase complexes to denaturing agents such as 7 M urea as the inhibitor will be inactivated and an active RNase will be released. Both inhibitors can be included in certain enzymatic reactions. However, RNasin is compatible with a wider variety of reactions conducted in vitro such as nuclear run-on transcription and translation in cellfree systems. The inhibitors are widely discussed throughout this volume.