Dear Nick, If you have access to a fluorescent microscope, you can try staining crystals with PicoGreen and seeing if they fluoresce. (see ref: Kettenberger H & Cramer P, 2006, Acta Cryst D v62 pp146-150: Kettenberger H, Cramer P.Fluorescence detection of nucleic acids and proteins in multi-component crystals.)
I've had good luck harvesting & dissolving crystals and then running a gel (stain with SYBR-Gold to increase sensitivity). If you can't get enough material to detect a band, you can radioactively kinase the dissolved crystals and then run a gel. Best Regards, Allyn > Dear all, > > I am trying to solve the structure of a transcription factor in complex > with its DNA. I got crystals of the complex under different conditions > than the protein alone and they also look different. Unfortunately, they > only diffract to 6Å so far. Before I continue to optimize the crystals I > would like to confirm that the crystals really contain the bound DNA. > Thus I tried to crystallize the protein alone under the same conditions > as the complex which did not give crystals. I also tried to stain the > crystals with methylene blue, but that did not work (but staining with > IZIT did not work either). Additionally I dissolved a crystal and > measured the absorption. The ratio between A260 and A280 was 1.3. So > there seems to be DNA, but less than there should be. > > Does any of you know a good way to quickly but reliably confirm the > presence of DNA in my crystals? > > Thanks a lot in advance. > > Nick > -- Allyn J. Schoeffler Berger Lab Dept. of Molecular and Cell Biology UC Berkeley phone: (510) 643-9491
