Sodium Acetate Precipitation of Small Nucleic Acids
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实验步骤
1. Add 2 μl carrier to the nucleic acid solution and mix well.
2. Add 1:10 volume of 3 M sodium acetate and mix thoroughly; for 230 μl of Lower Running Buffer, this will be 23 μl of 3 M sodium acetate.
3. Add 4 volumes of 100% ethanol; i.e. 1 ml 100% ethanol for 230 μl Lower Running Buffer plus 23 μl 3 M sodium acetate.
4. Mix thoroughly and incubate at –20° C overnight (16 hr).
5. We found that overnight incubation is important for maximizing recovery in this precipitation. (For example hr incubations resulted in significantly lower yield than overnight incubations).
6. Microcentrifuge at top speed for 30 min at 4° C or room temp.
7. Carefully remove and discard the supernatant.
8. Wash the pellet by adding 500 μl 80% cold ethanol. Microcentrifuge at top speed for 10 min at 4°C or room temp, and carefully remove and discard the 80% ethanol.
9. Repeat the 80% ethanol wash if needed to remove the large salt pellet from the sample.
10. To remove the last traces of ethanol, quickly re-spin the tube, and aspirate any residual fluid with a very fine tipped pipette or syringe needle. Air dry the pellet.
