[Combining approach with multiplex PCR and MLPA to detect deletion and duplication in DMD patients, carriers, and prenatal diagnosis].

OBJECTIVE

Applying multiplex PCR and multiplex ligation-dependent probe amplification (MLPA) in a clinical setting to detect deletions and duplications in the Duchenne/Becker muscular dystrophy (DMD/BMD) gene not only for patients, but also for identification of possible carriers and prenatal diagnosis.

METHODS

Multiplex PCR was used first in patients clinically diagnosed with DMD/BMD to examine 26 exons for a large deletion in the two hot regions of the dystrophin gene. For patients without a deletion detected in the aforementioned regions, MLPA was used to further examine all 79 exons to determine whether a deletion in the remaining non-hot regions or any duplication was present. A similar approach was applied to suspected carriers. In requested prenatal diagnosis cases, specific PCR was used to detect deletions, while MLPA was applied to detect duplications.

RESULTS

Multiplex PCR was used to examine 26 exons within the two hot regions in the Dystrophin gene for 22 patients with DMD; 13 (13/22) had multi-exon deletions. For the 9 patients without deletions in the 26 exons, MLPA was used to examine 79 exons. 3 patients had duplications, 1 patient had a single deletion in exon 18, and no deletions or duplications could be detected in the remaining 5 patients. Of the 16 carriers, 2 out of the 3 that had family history had deletions, while the other 13 carriers were mothers of affected children who were sporadic patients without family history. Of them, 8 mothers were carriers for either deletions or duplications. For prenatal diagnosis, 9 fetuses were examined (one case was twins). Of them, 2 fetuses had familial deletions or duplications detected. These results were verified after induced abortion. In 7 fetuses, no deletions or duplications were detected and all developed into children.

CONCLUSION

Multiplex PCR can detect 92.86% of deletions and is useful for prenatal diagnosis of deletions because it is simple, reliable and inexpensive. It can be the first choice in DMD/BMD gene diagnosis. MLPA is important for detecting deletions in non-hot regions/exons and duplications in the DMD/BMD gene, as well as for carrier detection.