Help to realize northern blot with non radioactive probe - (Feb/19/2014 )
I would like to realize a northern blot with a non radioactiove probe :
1- What do you think about biotin-labeled probe?
2- Do you use "Licor system"?
3- Which percent of unmodified dTTP and biotin-16-dUTP?
Thanks for your help!
I prefer a dig rather than a biotin probe. Roche is the primary (sole?) provider. I'd suggest 10-25% dig or biotin - UTP. Don't forget that you can't use most high fidelity PCR enzymes with dUTP -- Use straight Taq.
I don't know anything about the Licor system.
I'd strongly suggest that you start by detecting serial dilutions of your probe spotted on a membrane, then switch to serial dilutions of your target spotted on a membrane, and only when that is working well (low background, high sensitivity) should you worry about running gels and blotting.