Cross-linking T7 RNAP to T7 genomic DNA

[ramvenkat1983]

Hi...

I would like to cross-link T7 RNA Polymerase with T7 genomic DNA in-vitro. Both the RNAP and genomic DNA were purchased commercially. The interaction between the RNAP and T7 DNA are very weak. I would like to fix the proteins to the DNA to facilitate my experiments. Is there any method I can follow to do this? I have heard of formaldehyde cross-linking and UV cross-linking methods but I would like to have some specific protocol to try out.

Have a good day!!!

[admin]

Hi, no need to crosslink with formaldehyde UV-crosslinking is usually enough.

I would focus on buffer conditions, HEPES vs X Buffer; pH; Salt conditions; Mg conditions etc and optimize your binding first.

Cheers

[ramvenkat1983]

Thanks... What is the best method to check the results for the DNA-protein binding? Will gel electrophoresis give good results to make sure whether some binding reaction took place? I am concerned because the binding between T7 DNA and T7 RNAP is very weak.