urea/glycerol PAGE and cutting out the band... - cant find out protocols (Apr/24/2006 )
so I have this problem i'm breaking my head over, I just can't find the proper protocols for my purpose (s).....Breifly (for ones who dont already know ) I have expressed protein from E coli which has some degraded bands on SDS-PAGE...I want to isolate the upper band....since my protein is insoluble ...i have prepared it as inclusion body in 8M urea....now i want to use urea PAGE (with glycerol) to isolate the band and cut it out from the gel....then dialyse it.... ....my professor gave me some table of urea/glycerol PAGe buffers but it is so not clear and not complete.... ....and i cant find out any protocol to tell me how actually I dissolve this band out of the gel?!?! I am still searching and maybe will find out some protocols....will appreciate any help so so much....thank you!
...anyone?? and if possible do you think voltage settings should vary or same as SDS-PAGE?? ....