DNA length to be amplified in ChIP. - What should be DNA size range? (Aug/14/2005 )
what should be DNA length (range) to be amplified in ChIP assay?
whether ~1kb is feasible/ possible to amplify?
and what criterian to be considered while designing the primers, specially for ChIP?
Thank you in advance
I would amplify a length <= the average size of the sonicated gDNA sample. In most cases this is <=500bp I have seen primers used that amplify regions as small as 70bp.
You should design primers that span the TF binding site of interest, or if this is not possible, a region as close to that tfbs as possible.
You should follow general rules of primer design, and then I think it is especially important for ChIP to blast your primers to the genome of interest. (use the search for short, nearly exact matches function at NCBI blast.) It is most important that the 3' end not have any matches, I look for at least 2-3bp mismatch at the 3' end.
I suppose it is possible to amplify 1kb, as long as your average size for sonicaton is >= 1kb, but I wouldn't do so, probably better to sonicate the sample more and amplify smaller region...
hth, and good luck!