IGEM:Harvard/2006/DNA nanostructures/Notebook/2006-8-5

PAGE Coomassie Gel Imaging of Superconcentration of Eb, Fb, and Gb

 * destained: 10 min in H2O, two times
 * imaged gel from yesterday




 * Conclusions:
 * streptavidin band clear
 * biotinylated oligos coomassie band clear too, in different place
 * there are definitely streptavidin-bound biotinylated oligos in lane 3
 * it is difficult to see in the image, but a darker blue band was visible at the top of lane 5, where the nanostructure should be
 * streptavidin bound to Eb
 * unfortunately, Eb = inside-biotinylation -latches, so how the streptavidin bound and was visualized when the Gb (outside-biotinylation +latch1) didn't bind it is questionable
 * it is a very faint band
 * no bands were seen at the top of the gel in the expected lane 6 (Gb)


 * is it possible that streptavidin actually bound to nanostructures can't be stained w/ coomassie?
 * that would explain why the inside-biotinylated +latch1 design, which would not (we expect) permit the biotinylated-oligos within to bind streptavidin, shows a lot more unbound streptavidin and biotinylated-oligos-bound streptavidin

PAGE EtBr Gel Imaging of Superconcentration of Eb, Fb, and Gb

 * as a first try, the gel was destained a further 30min in H2O before EtBr staining was attempted
 * 200mL of H2O +7uL of EtBr were used as a stain; the gel was allowed to stain for 15 min


 * no luck, no bands at all were seen under UV
 * will save gel in H2O till figure out if there is anything to be done about it

More Folding Reactions for Further Superconcentration Experiments with Stains-All

 * made working stocks (=12.5 rxns) for:
 * outside-biotinylation w/o latches, Eb
 * outside-biotinylation w/ "future" (aka four-oligo, aka latch2, aka staple latches) latches, Ib
 * latching which can be attempted post-incubation
 * can be incubated at 37&deg;C for 1hr with the zipper oligos (c4.0.23, 24, 25), though latches can be used in the first incubation
 * inside-biotinylation w/o latches, Db
 * inside-biotinylation w/ latches, Hb


 * folded according to given protocol, using FOLDINGD

Folding for MgCl2 and 6x Titrations, Take Two

 * because the gel from yesterday of the previoius titration showed no bands whatsoever, the reactions were refolded as before
 * the reactions were, as usual, folded with the FOLDINGD program

3D models continued
Added scales to two images; Sketchup file now posted under Design 5 schematics.