Low Signal from Using N-ChIP for Weak DNA-Binding Proteins (ChIP (Low Signal))

Question

Accepted Answer

Consistently low or absent signal when studying transcription factors or chromatin-associated proteins using native ChIP (N-ChIP). Histone ChIP experiments work well in the same laboratory. Common causes: 1) Target proteins have weak DNA affinity and dissociate during N-ChIP 2) Proteins located far from DNA require cross-linking for stabilization 3) N-ChIP suitable only for tightly associated proteins like histones 4) Lack of formaldehyde cross-linking allows protein-DNA complex dissociation Solutions: 1) Use X-ChIP (cross-linked ChIP) for proteins with weaker DNA affinity 2) Use X-ChIP for proteins located far from DNA (e.g. chromatin remodelers) 3) Cross-link with 1% formaldehyde for 10-15 min before cell lysis 4) Reserve N-ChIP exclusively for histones and very tightly bound proteins