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Crude oligonucleotides can be precipitated after deprotection directly from the ammonia solution without the need for a separate step to remove the ammonia. This procedure removes small molecular weight impurities such as the ammonia itself and the by products of deptrotection (benzamide, isobutyramide). It is taken from a published method (M. Sawadogo and M.W. Van Dyke, NAR 19(3), 674 (1991).
1. The ammonia solution of the deprotected oligonucleotide (100ul) is mixed with 1 mL of n-butanol (ACS reagent grade) in an Eppendorf tube, is vortexed for 15 sec, then centrifuged for 1 min at 12K.
2. The H2O-containing butanol is removed and discarded. The pellet is redissolved in H20 (100ul) and re-extracted with butanol as above.
3. The final pellet (which may or may not be visible) is then dried under vacuum (desicator or lyophilizer) and resuspended in TE
4. The amount of oligonucleotide present should be determined at this stage - you will need to do a 1 in 30 to 1 in 50 dilution for this (from a 0.2 mmole scale synthesis). Typical yield (for a 25mer) is about 50 ug per 100ul of ammonia solution.