Problem with visibility on third comb in gels - (Jun/27/2012 )
Im using a mapping population to find a gene that gives resistance to a disease called stripe rust in a wheat variety. Currently I am running gels to test the polymorphism for the markers in the F2 population.
Im using 3 combs with 36 wells each on every gel I run. On several of the gels I run I can clearly see the DNA on first two combs when I take the picture under UV light but the DNA samples on the third and final comb are hardly visible.
Could this be a problem with the buffer or gel? Does anybody know what it could be? Any help or comments would be greatly appreciated.
I attached a picture so you can see what im talking about.
Because the dye (e.g. EB) moves the opposite way from your DNA, that is why the bands at the bottom are not visible anymore. There are ways of walking around: 1) don't use the last row of wells, or 2) after running stain the gel with EB; or 3) add EB to your buffer.
pcrman on Wed Jun 27 23:22:41 2012 said:
Because the dye (e.g. EB) moves the opposite way from your DNA, that is why the bands at the bottom are not visible anymore.
That's very interesting...since we always stain our gels after running them it was never an issue...
Im having another issue now with my electrophoresis. For some reason the DNA is not moving down the gels as fast as it should. Im having to run the gels for about 20 hours (instead of the normal 4-5 hours). Does anyone know what the reason is or could be?
Try new (fresh) buffer, try different tanks and different power sources.