Abstract
Objective: To explore a method to solve the issue of interference in fluorescence quantitative PCR non-specific amplification for gene detection. Method: A three-step method was used for amplification, and the quantitative fluorescence signal collection process was set in the extension stage. Results: Three-step amplification has the advantages of wide application range; improved accuracy; and reduced primer design requirements. Conclusion: The interference of non-specific amplification signals was effectively avoided, the melting curve plotting process was omitted, the reaction time was shortened, and the detection accuracy was improved.