I'm working on proteasomal activity on C.elegans. I extracted worms with lysis buffer containing :20mM tris, 5mM MgCl2, 1mM DTT, 1mM ATP, 10% glycerol, 0,05 NP40. I ran a native gel electrophoresis (Tris-borat pH8,2) and I obtained this profile (see picture). The last well is empty of protein extract (right), I just put lysis buffer + xylene cyanol (traces) inside. I don't know if the waves that we see are due to detergent? Any suggestion ? thank you
Why are you using Tris-borate? This is for nucleic acids. The running buffer for native gels is normally Tris-glycine.
Hello, because I found several papers about proteasome using tris borate.. Do you think it could be because of that? It is very heterogenous, sometimes I don't have these waves and sometimes I have... I was thinking that maybe it's du to the separation ?
I do not like the way these samples seem to run round parts of the gel. Could this be that the gel is polymerising unevenly . Do you have fresh APS made up for each gel. If you only sometimes get these results perhaps you should polymerise for longer and always with fresh APS and mix the gel constituents well before casting the gel
- Badges
- Science method