Hi,
I want to make a 10% SDS-PAGE gel that contains 6M urea. Both the resolving and stacking buffers should have urea. Could anybody please suggest how to do this? I prepared my samples in buffer containing Tris-Cl, SDS, glycerol and bromophenol blue followed by heating at 100C.
I want to make a 10% resolving gel and 5% stacking gel both containing 6M urea.
Please advise.
Thanks
Sumit
Just caluculate the needed amount of Urea for the final volume of gel solution. Be carefull to dissolve slowly and don not heat up the solution to high for quicker dissolvment. As the volume of your solution will increase , make sure to add the water at last to adjust the final volume. and prepare an new loading buffer, which also contains 6 M Urea. This will sharper your bands if you use the same concentrations in bith gel and loading buffer.
Here is a publication which might be helpful:
Article Denaturing Urea Polyacrylamide Gel Electrophoresis (Urea PAGE)
- Badges
- Science method