Hi, I am trying to crystallize a protein-DNA complex. I purify the protein finally using gel filtration. I purchase single stranded complementary oligos (desalting from idtdna.com), mix them up and make DNA duplex by heating to 95 degree C and cooling to room temperature. I mix protein and DNA, concentrate and use it for crystallization. I am geting small crystals consistently under a specific condition. These crystals take up IZIT dye but are not well shaped. I am not able to improve the size and shape of the crystals substantially even after screening with additives (Hampton research). I suspect that purity of the duplex DNA (presence of unpaired oligos) is limiting the chances of obtaining better crystals.
How can I purify the duplex DNA further? Are there better ways of making protein-DNA complex for crystallization? If I make the protein DNA complex and then do the gelfiltration, will the complex purified so be a better choice for crystallization? Thank you Kumar Dept. of Biochemistry, Cellular and Molecular Biology, Walters Life Science, # 406, University of Tennessee, TN, Knoxville, USA
