Detection of amplified oncogenes by differential polymerase chain reaction

Abstract

Oncogene amplification has been found in a variety of human cancers and may have prognostic importance. Therefore, techniques which facilitate detection of gene amplification could have wide applicability. We have devised a sensitive, rapid, and non-radioactive procedure for detecting alterations in gene copy number based on the polymerase chain reaction (PCR). In this technique, called differential PCR, a target gene and a single-copy reference gene are co-amplified by PCR in the same reaction vessel. The level of target gene amplification is reflected in the ratio between the two resulting PCR-product bands. We show that this method can detect as low as two-fold amplification of specific target genes. Furthermore, amplification of neu and the epidermal growth factor receptor gene could be detected in as few as 100 breast carcinoma cells or in single sections of formalin-fixed, embedded material.