CTab procedure is not working, results in a milky solution. - (Sep/08/2014 )

Hello friends.

We have a hard problem with CTab procedure in plants. We usually extract DNA from Oaks, Pines, Platanus spp., and Oregano (Polyominthia spp.) and like 1-2 months ago the final product its a "milky solution" when its freezed at -20ºC. The problem doesn't stop here, because when we charge the "milky" samples to an agarose gel 1% it doesn't run, and of course doesn't work at all. We use this protocol for like 7 or 8 years ( Doyle & Doyle protocol), so we already had good results with all type of plants we usually use, and right now is not working.

So, we made new CTab buffer and changed all the reagents and we still have the same issue.

Any suggestions?

I added an image of few samples we had, they are freezed and when you unfreeze them, they are transparent.

Thank you.

Alex.

My guess is that you have either not extracted the DNA in a pure enough form so that you are getting polysaccharides coming through the extraction (too much input material?) or that someone is carrying over some of the phenol layer into the subsequent extraction.

Usually the ctab buffer dissolves only under "high" temperature (> 45). Normally it has required to be at 65 degrees in DNA/RNA extraction protocols.