The development of preimplantation genetic diagnosis for myotonic dystrophy using multiplex fluorescent polymerase chain reaction and its clinical application

Preimplantation genetic diagnoses (PGD) for single gene defects require considerable time and resources for the standardization of polymerase chain reactions that are rapid, sensitive and reliable. Developing tests for the trinucleotide repeat diseases, where the expansion of unstable repeats produces the phenotypes, are particularly complex. One of these disorders is myotonic dystrophy where, at present, diagnosis at the single cell level relies on the detection of the normal alleles from both the affected and unaffected parent. The incorporation of short tandem repeat polymorphisms in the assay can give additional information to improve the accuracy of diagnosis. We have developed a multiplex fluorescent reaction for myotonic dystrophy and one of two closely mapped, highly heterozygous, short tandem repeats (D19S219 and D19S559) on chromosome 19 to reduce the possibility of misdiagnosis due to contamination, act as a control for allelic drop-out and maximize the number of embryos genotyped. This protocol was designed as a general diagnosis for myotonic dystrophy, using the most informative of the two polymorphisms for each couple. Subsequently this approach was used in a PGD treatment cycle.