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Fluorescent PCR: a new technique for PGD of sex and single-gene defects
1Institute of Epidemiology and Health Services Researh, University of Leeds, UK. ianf@epid.leeds.ac.uk.
Background:
For single-cell diagnosis, particularly preimplantation genetic diagnosis to be successful four main criteria must be achieved: sensitivity, reliability, accuracy, and identification/elimination of contamination.
Methods And Results:
Fluorescent PCR achieves all four necessary criteria and, in addition, currently allows genes on up to nine chromosomes to be simultaneously investigated. Fluorescent PCR has high sensitivity (approximately 1000 x conventional analysis systems), high reliability (97%), and high accuracy (97%) rates for both sex and CF diagnosis in single somatic cells. The low detection threshold allows allelic dropout (one of the main causes of misdiagnosis) to be easily distinguished from PCR phenomena such as preferential amplification. High reliability (90%) and accuracy (97-100%) have been achieved in sex and CF diagnosis in human blastomeres. Fluorescent PCR can also be used to DNA fingerprint (STR profiling) single cells to identify the source/origin of the cell and determine if contamination has occurred.
Conclusions:
Fluorescent PCR is therefore a suitable method for PGD.