High Signal in All Samples Including No Template Control (ChIP (PCR Amplification Problems))

Question

Accepted Answer

PCR amplification occurs in all samples including no template controls (NTC), producing Ct values or visible bands in negative controls. This indicates systemic contamination rather than specific ChIP enrichment. Common causes: 1) Real-time PCR master mix or buffer solutions contaminated with template DNA 2) Contaminated primer stocks from previous experiments 3) Aerosol contamination from previous PCR reactions in the preparation area 4) Pipettes or tubes contaminated with amplicon carryover Solutions: 1) Prepare fresh real-time PCR solutions from stock reagents 2) Use new aliquots of primers and verify NTC is negative before proceeding 3) Physically separate PCR setup area from post-PCR product handling areas 4) Use dedicated pipettes and filter tips for PCR setup 5) UV-treat PCR setup area and clean surfaces with DNA decontamination solution