Asymmetric pcr amplification, its special primer and application

Abstract

The present invention discloses an asymmetric PCR amplification method, its special primer and application, aims to provide a simple, effective PCR amplification for preparation of single-stranded product. The asymmetric PCR primer of the invention comprises some PCR primer pairs, in which an unrelated nucleic acids sequence to target sequence to be detected is added onto 5'-terminal of one primer. The asymmetric PCR amplification provided includes the steps: 1) preparative denaturing; 2) repetitiously denaturing, primers annealing, extending cycles as the first stage of PCR amplification; 3) repetitiously denaturing, primer extending cycles as the second stage of PCR amplification, wherein an unrelated nucleic acids sequence to target sequence to be detected is added onto 5′-terminal of one PCR primer of each pair in extension. With the asymmetric PCR amplification of the invention, high throughput of single-stranded products can be obtained, single PCR amplification or multiple PCR amplification can be carried out. And the method can be widely used in detection of nucleic acids.