Shawn and William suggested that for an initial assay, it'd be easy to just stain protein with Coomassie blue. So that's what we attempted.
We ran two 10-20% native polyacrylamide gels at 15 V overnight; the first used some lanes from the Nano group's gel. Visualization to come tomorrow.
The composition of what we added to each line is below; materials were added and incubated for 30 minutes in 2 uL total volume for lanes 1-11, 1-12 prior to dilution to 10 uL including addition of loading dye. Lanes 2-1 to 2-9 were incubated for an hour; longer incubation carried out since DNA basepairing reactions for lanes 2-10 to 2-12 were carried out for 30 minutes using results from 30-minute incubations of lanes 2-1 to 2-9. This incubation time should not be significant.
Materials used: Thrombin: a 2 uM solution of Thrombin. Streptavidin: a 2 uM solution of Streptavidin. Thromb5, Thromb20, Thromb35, Thromb50, Strep5, Strep20, Strep35, Strep50 are described in Plan section above. Bock's buffer is a reaction buffer for DNA-protein binding, used in several papers (see Nano group's page). 10X dye is a few flakes of bromophenol blue + 500 uL Bock's + 500 uL glycerol.
All quantities in uL.
|4X Bock's Buffer (see Nanostructure page)||.5||.5||.5||1||1||1||.5||.5||1||1||1|
|Bock's 1X buffer||7||7||7||7||7||7||7||7||7||7||7||5||5||5|
Ran 2 10-20% polyacrylamide gels along with nano group.