Effect of unexpected sequence interruptions to conventional PCR and repeat primed PCR in myotonic dystrophy type 1 testing.

Myotonic dystrophy type 1 (DM1) is caused by expansion of the CTG trinucleotide repeat in the DMPK gene. Our study focuses on the effect of recently described unusual sequence interruptions inside the CTG tract on conventional polymerase chain reaction (PCR) and triplet repeat primed PCR (TP-PCR) amplifications, which are the methods now widely used in molecular testing for DM1. For molecular characterization of the CTG repeat tract, we used conventional fluorescent PCR with bidirectional labeling and both forward and reverse direction TP-PCR. Though the results of the methods are still unambiguous for most alleles, mistyping and false results may occur in the typing of some unordinary alleles carrying sequence interruptions. The presence of these interruptions may lead not only to altered TP-PCR profiles, as can be expected, but also to abnormal electrophoretic mobility of complementary strands produced by conventional amplification of such alleles. Our findings suggest that the simultaneous combination of bidirectionally labeled conventional PCR with TP-PCR performed in both directions may be necessary for increasing the reliability and accuracy of the TP-PCR-based assay for DM1 testing.