A successful strategy for preimplantation genetic diagnosis of myotonic dystrophy using multiplex fluorescent PCR

The most common form of inherited muscular dystrophy in adults is myotonic dystrophy (DM), an autosomal-dominant disease caused by the expansion of an unstable CTG repeat sequence in the 3' untranslated region of the myotonin protein kinase (DMPK) gene. Expanded (mutant) CTG repeat sequences are refractory to conventional PCR, but alleles with a number of repeats within the normal range can be readily amplified and detected. Preimplantation genetic diagnosis (PGD) of DM has been successfully applied. However, a misdiagnosis using the reported protocol was recently documented. Two new PGD protocols for DM have been developed which utilise multiplex fluorescent PCR. Ideally a linked polymorphic marker, APOC2, is amplified in addition to the normal DMPK alleles, thus providing a back-up diagnostic result. However, the two couples reported in the present study were not fully informative at the APOC2 locus and so an unlinked short tandem repeat (STR) marker, D21S1414, was substituted. The highly polymorphic nature of the D21S1414, DMPK and APOC2 loci means that a very simple genetic fingerprint can be generated by analyses of these loci. This allows most DNA contaminants to be detected. Contamination is a significant problem for PGD and is the primary reason for the inclusion of D21S1414 and APOC2 in this protocol. This paper reports the first clinical experience and pregnancies following PGD for DM using a multiplex fluorescent PCR protocol.