Posted 28 January 2010 - 01:45 PM
I'm trying to figure out how to design a northern blot, I have DNA in Topo TA cloning vector with T7 promoter so what is the next step should I use Prime a Gene to label it and I don't really understand the non label - how do you detect the hybridization?
Thanks for helping
Posted 29 January 2010 - 09:42 AM
and another thing is this system preferred to the fluoresence or the biotin?
Posted 31 January 2010 - 03:17 PM
If you use the luminescence method (alkaline phosphatase with CDP) then you need standard radiographic film (I use kodak x-omat bt) or a camera set up for taking pictures of luminescence. You can also detect using chromogenic substrates (NBT/BCIP) which will give a coloured band on your membrane, but is slightly less sensitive than luminescence. I haven't used a fluorescence method, so I can't comment.
Posted 03 February 2010 - 09:51 AM
I decided to use the biotin as nonisotopic labeling and I bought the whole system from ABI with the MAXIscript and so on but now I found out that they have template for housekeeping genes (GAPDH, b-actin) only for mice and not for human. Do you know if there are commertial templates for human or do I need to design myself?