For all the DNA gurus....help with "invisible" pellet - Problematic EtOH precipitation (Jan/09/2006 )
I see how everybody is handing neat advices about EtOH precipitation of DNA , but...when aspirating the supernatant after centrifuging your DNA + EtOH+salt, how do you aspirate correctly when you DON'T SEE the pellet!?!? In my lab, I am not "allowed" to use carriers because in both of my PI's hands and my senior the method of old school EtOH precipitation of DNA works, so my task is to make it working in my hands...But my hands seem to be clumsy!!! Whenever I don't see my pellet after centrifugation, I loose it by aspirating:(. Help somebody, pleeeease!
be careful about the way you insert your eppis.... the attached .doc is german, but still you'll get the meaning. if you insert the eppis like that, the dna will end up at the "outer" eppi wall, that is, the eppi wall most away from the center. so be sure to place your eppis in a way you'll remember were to find the precepitate back 
in the picture, red arrow marks the spot
cheers, k.
in the picture, red arrow marks the spot
cheers, k.
Thanks for sharing, I did learn that however a half a year ago:D! I think it's my pipeting that's problematic, I probably jerk the tip of the pipette in the solution by fast movements and just lose the pellet in the end. What happens is that the "pellet", if I see it of course, gets loose from the tube wall the second I enter last 5 microliters of the supernatant and then it gets dissolved. It got me thinking that why sequencing precipitations work usually good is becasue of the 70% EtOH wash, but hey, guess who is not "allowed" to useit in the "normal" ppt.!?
Take care!
I never aspirate my ethanol after precipitation/centrifugation, but decant it in one swift movement. Pipetting often disturbs the pellet, but simply inverting the tube and allowing to drain does not
usually pellet sticks well after EtOH precipitation, but you have t handle it carefully.
usually it de-stick when you stop aspirating with your pipet, then you should avoid to aspirate the little drop which stays after the first aspiration.
(not easy to explain in English) : don't re-pipet in you tube.
you can also do as John says. I usually do it also, and I absorb the last drop with a paper (never put the paper inside the tube, don't let the last drop go back to the bottom of the tube, it will unstick the pellet. however if you did, don't aspirate the last drop, don't absorb it on a paper, let it inside the tube with the floating pellet and dry a little longer.
Hi, when you ppt DNA using isopropanol generally the pellet will be invisible. Though I generally just decant the tube and dry the pellet. Even if the pellet is invisible asume the presence and gently decant.