Image analysis in quantitative PCR. An application for the measurement of c-erbB-2 oncogene amplification in DNA from human tumours
✍ Scribed by Orlando, C. ;Sestini, R. ;Zentilin, L. ;Gelmini, S. ;Pinzani, P. ;Giacca, M. ;Pazzagli, M.
- Publisher
- John Wiley and Sons
- Year
- 1994
- Weight
- 511 KB
- Volume
- 9
- Category
- Article
- ISSN
- 0884-3996
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✦ Synopsis
We present a n application o f image analysis for the direct quantification o f PCR products after gel electrophoresis and ethidium bromide staining o f DNA. This procedure has been applied t o the development of a n assay based on competitive PCR for the measurement o f the degree o f amplification of c-erbB-2 oncogene in DNA from human tumours. In this method t w o DNAspecies (genomic and competitor) compete for PCR amplification. Since results are calculated from the final competitor/ genomic ratio any variable affecting the rate o f PCR amplification has no effect on the accuracy o f the ratio measurement. Results are reported which show that even large variations in the experimental conditions (number of PCR cycles, sample volumes and extracted DNA quality) did not interfere w i t h the precision of the measurement o f the competitor/genomic ratio.