starting amount of DNA for ChIPs - using the ChIP for ChIP-on-chip analysis (Apr/11/2006 )
Can anyone of you suggest, what starting amount of DNA I should use for a reproducible ChIP-assay?
I found different protocols talking about 100 to even 300 µg per ChIP. Is that really the common way to conduct a ChIP? And why do I have to use so much DNA? I've been told, that the normal range of precipitated DNA is around some ng, so where do I loose so much DNA? Is there any possibility to start with lower amounts and still get good chipping results?
My problem is, that I want to analyse primary cells and therefore I don't have so much DNA 'available'....
Thanks already in advance for your help!!
The important thing here is not the amount of DNA but the protein (histone, or whatever) that is associated with the DNA. After immunoprecipitation and washing, anything that is not bound by the antibody is washed away and the remaining DNA may be too little to give good signal in PCR amplification.
I don't mesaure how much DNA I use for ChIP but use a certain amount of cells, such as 1x10^6.