Mutation detection in the DMD gene has historically been challenging, due
mainly to the large size of the gene and the fact that over 90% of the point
mutations are unique. Although hot spots have been reported for exon deletions
and duplications no such spot has been identified for point mutations which are
scattered across the whole gene.
Materials and Methods
In our study we present a general overview of the molecular analysis findings
in 645 patients (608 male and 37 female) referred to our department during the
past 2 decades for DMD gene testing. Exon deletions and duplications were
identified using either the standard 18 exon multiplex PCR (mPCR) or the MLPA
method while Enzymatic Cleavage Mismatch Analysis protocol (Vogiatzakis et al,
2007; 2010) followed by direct DNA sequencing was performed for the detection
of point mutations.
The deletion boundaries, where not apparent, were further investigated with
either mPCR or MLPA.
The study revealed 309 alterations comprising 254 exon deletions, 24 exon
duplications and 31 point mutations. Several polymorphisms were also identified
on the DMD gene, of which 6 single nucleotide alterations remain still under
investigation regarding their possible pathogenic character. Although the
majority of the mutations were detected along the DMD gene hotspot regions (as
mentioned in literature), minor discrepancies were noted concerning the
boundaries of the exons’ deletions.
1. A non-random distribution concerning a high incidence of deletions in the
distal hot spot (exons 45-55) and duplications in exons 2-8 was noticed and is
consistent with the results reported for previous large cohorts
(Tuffery-Giraud S et al, 2009, Aartsma-Rus A et al, 2006). Among the 31 point
mutations disclosed, a rather high detection rate was noted in exons 21 and 66
having 3 different mutations each.
2. This study revealed 31 point mutations on the DMD gene. Specifically, 37%
frameshift mutation 17% splice site mutation and 46% stop codon mutation. All
the results are in accordace to the international literature.
3.Mutations in the DMD gene should be studied even in individuals with chronic
subclinical CK elevations such as DMD carriers
4.It should be noted that 251 of the patients (males and females) were referred
solely due to minor BMD-like phenotypic symptoms such as hyperCKemia thus
explaining the reduced (47%) detection rate among referrals.
Dystrophin, ECMA, MLPA, Novel methodologies, DMD/BMD