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Open Access Highly Accessed Methodology article

A novel technique for measuring variations in DNA copy-number: competitive genomic polymerase chain reaction

Kyoko Iwao-Koizumi1, Kazunori Maekawa1, Yohko Nakamura2, Sakae Saito1, Shoko Kawamoto1, Akira Nakagawara2 and Kikuya Kato1*

Author Affiliations

1 Research Institute, Osaka Medical Center for Cancer and Cardiovascular Diseases, 1-3-2 Nakamichi, Higashinari-ku, Osaka 537-8511, Japan

2 Division of Biochemistry, Chiba Cancer Center Research Institute, Chiba 260-8717, Japan

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BMC Genomics 2007, 8:206  doi:10.1186/1471-2164-8-206

Published: 2 July 2007

Abstract

Background

Changes in genomic copy number occur in many human diseases including cancer. Characterization of these changes is important for both basic understanding and diagnosis of these diseases. Microarrays have recently become the standard technique and are commercially available. However, it is useful to have an affordable technique to complement them.

Results

We describe a novel polymerase chain reaction (PCR)-based technique, termed competitive genomic PCR (CGP). The main characteristic of CGP is that different adaptors are added to the sample and control genomic DNAs after appropriate restriction enzyme digestion. These adaptor-supplemented DNAs are subjected to competitive PCR using an adaptor-primer and a locus-specific primer. The amplified products are then separated according to size differences between the adaptors. CGP eliminates the tedious steps inherent in quantitative PCR and achieves moderate throughput. Assays with different X chromosome numbers showed that it can provide accurate quantification. High-resolution analysis of neuroblastoma cell lines around the MYCN locus revealed novel junctions for amplification, which were not detected by a commercial array.

Conclusion

CGP is a moderate throughput technique for analyzing changes in genomic copy numbers. Because CGP can measure any genomic locus using PCR primers, it is especially useful for detailed analysis of a genomic region of interest.