Precipitation of long ssDNA oligos to remove potential contaminants.
1.
2.
3.
4.
5.
6.
7.
8.
Resuspend the lyophilized DNA pellet from the vendor in 400 µl water.
Add 40 µl 3M Na-Acetate pH 5.2 and 1.0 ml 100% ethanol.
Incubate at -20˚ for 30 minutes.
Spin 15’ in microcentrifuge at full speed.
Carefully remove and discard the supernatant.
Add 500 µl 70% ethanol. Sping 5’. Discard the supernatant.
Repeat step 6 one more time.
Carefully remove all “dregs” of ethanol from the pellet and let it air dry 5
minutes.
9. Resuspend the oligo in embryo-grade water at an expected concentration of
about 1000 ng/µl based on the total amount of shipped oligo (check the
vendor’s info sheet that came with the oligo).
10. Measure by nanodrop. The 260/230 ratio should be >2.0.
DNA oligonucleotides are sometimes contaminated with unknown materials when
they arrive direct from the manufacturer. Although requesting additional
purification by the vendor may help this (e.g. PAGE purification), remember that
these purifications are usually designed primarily to increase the proportion of the
product that is full-length, and not necessarily to remove all potentially toxic
contaminants.
Here is an example of how DNA oligos may ship with non-DNA impurities, and that
ethanol precipitation can help remove these.
DPM 9/21/15