COLD-PCR: Improving the Sensitivity of Molecular Diagnostics Assays

Overview

Journal Expert Rev Mol Diagn

Specialty Molecular Biology

Date 2011 Mar 17

PMID 21405967

Citations 27

Authors

Coren A Milbury

Jin Li

PingFang Liu

G Mike Makrigiorgos

Affiliations

Soon will be listed here.

Abstract

The detection of low-abundance DNA variants or mutations is of particular interest to medical diagnostics, individualized patient treatment and cancer prognosis; however, detection sensitivity for low-abundance variants is a pronounced limitation of most currently available molecular assays. We have recently developed coamplification at lower denaturation temperature-PCR (COLD-PCR) to resolve this limitation. This novel form of PCR selectively amplifies low-abundance DNA variants from mixtures of wild-type and mutant-containing (or variant-containing) sequences, irrespective of the mutation type or position on the amplicon, by using a critical denaturation temperature. The use of a lower denaturation temperature in COLD-PCR results in selective denaturation of amplicons with mutation-containing molecules within wild-type mutant heteroduplexes or with a lower melting temperature. COLD-PCR can be used in lieu of conventional PCR in several molecular applications, thus enriching the mutant fraction and improving the sensitivity of downstream mutation detection by up to 100-fold.

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