EMSA - probe design and competition
Posted 12 July 2010 - 07:43 AM
The lab that I am working in is really new to EMSA and functional genomics as a whole. Part of my thesis project is to design an EMSA study for a particular SNP in the TAGAP gene. I've got protocols for EMSA both radiolabelled and non-radiolabelled. I'm having trouble finding advice on how to design my probes.
How long should my probe be? Is this dependent on how big the presumed transcription factor is?
Also - how do I design competitor probes? How many competitor probes do I need? Is there a protocol somewhere that will tell me this?
I appreciate any and all advice.
Posted 12 July 2010 - 06:50 PM
Posted 13 July 2010 - 08:22 AM
Posted 13 July 2010 - 09:31 AM
So does the sequences for the nonspecific competitor need to include anything specific (like a TATA sequence or a certain percentage of GC)- or is it just random sequence?
Posted 14 July 2010 - 07:03 AM
It doesn't have to, it can be random. In my case, I used sequences that contained binding sites for other, unrelated transcription factors, simply because I had them already in the lab.
Posted 18 November 2010 - 10:18 AM