I can't believe I messed this up
I was doing a DNA extraction with phenol chloroform. The protocol we use in lab has us use phenol, then phenol:chloroform, then chloroform. When I got the phenol:chloroform step, however, I used a bad bottle, which only had one phase (the wrong one, apparently, although I still don't understand where the right phase had gone). I noticed that after adding it, it didn't separate into two layers (organic and aqueous, as it is supposed to). So I centrifuged it. No dice. So I took the whole volume and added the right phenol:chloroform to it, and continued with the rest of the extraction. This involves using isopropanol, then two washes with 70% ethanol. Surprise surprise, no pellet precipitated out after using the isopropanol. So I spun it down anyway, got rid of the isopropanol, and then added 70% ethanol and stuck it in the -20 overnight.
I came in yesterday, spun it down, found a pellet (yay!) and then decided to wash it again with 70% ethanol. After adding the ethanol and spinning it down, the pellet disappeared. I put it in the freezer again, and this time, there was no pellet after spinning it down.
Where has the DNA gone? I didn't throw out the ethanol from the second wash, so it should still be there, right? How do I get it to form a pellet again?
Any light any of you could shed on this matter would be greatly appreciated.
