hi,

i think you must use the dilution for your samples, then perform your q

pcr analysis.

with the best regards

Hi Fabrizio,

Your IgG control is there to control for background levels such as nonspecifically pulled down chromatin. So, you need to use the same dilution and volume as input for both IgG ctrl and specific Ab samples when you compare them by qPCR. If you were to adjust for the DNA concentration in stead you would potentially get very different background measurements between your samples, which would hinder your comparison.

Best wishes,

Maarten

Thank you.

So basically the input is 50 microliter from 10 ml of chromatin. IgG and specific Ab samples comes from 10 ml of chromatin each. All of the DNA samples were resupendend in 40 microliter of H20.

Hi, you can't do this with a single qPCR. You need two loci and calculate relative enrichment with the delta Ct method since between-tube differences can be very high in ChIP, especially when you start doing ChIP.

Chapter Native Chromatin Immunoprecipitation