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Proc Natl Acad Sci U S A. 1999 Aug 3;96(16):9236-41.

Digital PCR.

Author information

1
The Howard Hughes Medical Institute and the Johns Hopkins Oncology Center, Baltimore, MD 21231, USA. vogelbe@welchlink.welch.jhu.edu

Abstract

The identification of predefined mutations expected to be present in a minor fraction of a cell population is important for a variety of basic research and clinical applications. Here, we describe an approach for transforming the exponential, analog nature of the PCR into a linear, digital signal suitable for this purpose. Single molecules are isolated by dilution and individually amplified by PCR; each product is then analyzed separately for the presence of mutations by using fluorescent probes. The feasibility of the approach is demonstrated through the detection of a mutant ras oncogene in the stool of patients with colorectal cancer. The process provides a reliable and quantitative measure of the proportion of variant sequences within a DNA sample.

PMID:
10430926
PMCID:
PMC17763
DOI:
10.1073/pnas.96.16.9236
[Indexed for MEDLINE]
Free PMC Article

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