This is my first post and I am desparately looking for some help recovering DNA from CsCl preps. I have recently run into some problems precipitating DNA using sodium acetate (3M 1:30 dilution) and 2.5 volumes of ethanol. I know the plasmid DNA is present as I remove it from tube post spin and then remove ethidium bromide with salt saturated butanol, I usually dialyse o/n in TE then recover the sample and complete the precipitation, but for some reason I have been unable to precipitate my DNA out of the last few samples I have run and I am not sure why, I know the DNA is there but after adding more and more ethanol the sample gets two large in volume and is cumbersome the handle, I attempted to to run the enitre sample over a qiagen maxi column and was able to recover some DNA but not at the concentration I am used to getting with the standard protocol. I also attempted to forgo the dialysis and add 3 vol H20 and 8 volumes ethanol and still nothing percipitated, even after incubation o/n at 4 degrees when I just added 1:10 volume sodium acetate and 2.5 volume ethanol a tremendous amount of salt precipitated, and now I dont know where the DNA is, if it is caught up in the salt precipitate or still in solution??? HELPPPP I really need this DNA and dont want to start another CsCL prep!!!
Edited by zimmersk, 14 June 2004 - 07:57 AM.