IGEM:Harvard/2006/DNA nanostructures/Notebook/2006-7-25

Oligos Order Form
c5.0 Partial Oligo Order Form

PEG precipitation

 * Used samples of 3.2.H and 3.2.I folded yesterday
 * 20 of each, 190 mL water, 50 mL 20% PEG / 2.5 M NaCl in respective 500  PCR tubes
 * gel analysis :
 * lanes 3 and 6 (reconstituted precipitate of H and I, respectively) appear to be free of oligos (compare to lanes 2 and 5, which are raw folding reaction), but yields are still very low, as evidenced by amount of nanostructures in supernatant (lanes 4 and 7)

Gel purification

 * 20 mL each of 3.2.D and 3.2.E run on a gel
 * nanostructure bands isolated and purified using Qiagen gel purification kit (good for fragments up to 10 kb according to manufacturer)
 * will run on agarose gel tomorrow to determine purity

Silver staining experiments

 * goal: to determine the minimum concentration of streptavidin that can be visualized with silver stain, and to note the times at which different concentrations become visible
 * methods: 12% PA gel with lanes listed below, run for 20 min. at 130 V, then stained with Bio-Rad Silver Stain Plus Kit and protocol.
 * 35 min. in fixative solution
 * 2 x 10 min. wash in water
 * XX min. in staining solution
 * 15 min. in stopping solution


 * results:
 * first three lanes (40k, 20k, 10k pmol) stain reliably
 * fourth and fifth lane lane (5k, 2.5k pmol) barely stains
 * the rest of the lanes (1.25k pmol and less) do not stain more than the background
 * this is disappointing (we can image 1 pmol of streptavidin with Coomassie blue)