Hi all,
I'm currently trying to separate a couple of dsDNA fragments (160 and 170bp)
The DNA itself smiles as the gel run making the separation two fragments difficult. Are there any suggestions to reduce this effect?
The electrophoresis condition I've used are:
Bio-Rad Mini Protean Gel Tank. 1.0mm Glass plate. 10 well comb. Set gel for 2h, 12% polyacrylamide, 1.0x TBE, 80V, 3h.
Thank you
reduce the voltage and run for a bit longer. Load empty lanes beside the sample with loading dye to ensure that the lanes each side are running the same as your samples. smiling is usually caused by uneven heating of the gel so even thickness of the gel and heat dissipation ( often with a metal plate on sone systems) is important
@Paul: The gel overall smiles very little, it's the sample within the well. The bands curve.
yes but there is uneven heating within lanes due to different salt concentrations changing the electrical resistance of each track so even tracks can run at different temperatures so loading blanks with dye either side of the sample often evens out the heating effect across the track works if you do not already have a full gel. be careful also to flush out the wells prior to loading and possibly load less sample if you have a good signal
- Badges
- Science method
- Similar topics
- Biological Science
- Ecology