- Modified oligonucleotides are also dissolved in sterile water at pH 7,0 or in one of the named buffers. Cyanine dyes are instable in alkaline environment. Particularly for Cyanine5-modified oligonucleotides one should take care that the solvent really has a pH of 7,0 !
- Oligonucleotides are best stored in dry state. Therefore, they should only be dissolved only immediately before use.
- Appropriate working conditions and proper handling in a nuclease-free environment ensure a long life for RNA and DNA. This should be preferred even to DEPC-treatment (see below).
- The use of EDTA (e.g. in Tris) does not influence the oligonucleotide itself, but should be taken into account with regard to subsequent enzymatic reactions.
- The use of water with a pH below 7,0 leads to depurination of the oligonucleotide. Use NaOH to increase the pH or dissolve the oligonucleotides in a suitable buffer.
- DEPC (diethyl-pyrocarbonate) treated solvents are, even in combination with following RNase-sensitive methods, not recommended due to the high toxicity. Besides a moderate genotoxicity of DEPC itself, the highly carcinogenic ethylcarbamate is formed when combined with ammonium ions, as are used e.g. in the precipitation of nucleic acids. Further DEPC modifies purines in single strands. RNA that is modified in this way will not be reverse transcribed properly (Ehrenberg, Fedorcsak and Solomosy, Prog Nucleic Acid Res Mol Biol. 1976;16:189-262, Mandiyan and Boublik, 1990, NAR 18, 7055-7062).