Hi Annarita,

This might help you,

Primers for a locus where you know your protein or modification of interest is present (positive control locus) and one where it is absent or significantly decreased (negative control locus) should both be tested in RT-PCR. This will tell you whether the observed enrichment is specific. It is important to include these controls as some antibodies result in non-specific enrichment.

Gud luck.

 As Amar said both, positive and negative controls are important. Depending on the modification that you are checking you need to look for the specific control. For repressive marks I use very active genes as negative control, for marks enriched in the coding region I use intergenic ones and so forth.

The ENCODE datasets can provide some pointers to negative and positive controls for a given cell type and histone mark.

Both positive and negative controls are usually required. The problem with negative target controls is that the signal may be very low and somewhat variable. No way round this except to have multiple negative controls.

Hi Annarita,

Did you carry out your ChIP experiment using IgG antibody (negative control) and H3 antibody (positive control)? Sometimes it's very difficult to know which DNA region it's the best option for negative controls, but you could compare your target signal with the inmunoprecipitation from negative and positive antibodies. Also, in the qPCR you can normalize with the CT of the input.

For histone ChIPs it is very easy to find a negative control region but it really depends upon the Hmod you are trying to ChIP. More information would help. If you mean negative regions devoid of active marks (i.e. inactive and not transcribed) then good examples could be PRSS1 or KRT5 (two genes only expressed in specialised cells) TSS or intergenic regions.