Diagnosis of Duchenne and Becker muscular dystrophies by polymerase chain reaction. A multicenter study
OBJECTIVE--To assess the efficiency, reliability, and ease of use of DNA
diagnosis for Duchenne and Becker muscular dystrophies (DMD/BMD) using the
polymerase chain reaction (PCR). DESIGN--DNA from the patients was screened
for deletion mutations using multiplex PCR, and the results were compared
with those obtained by Southern blot analysis. The PCR multiplex reaction
detects nine specific "hot-spot" exons in the dystrophin gene while the
Southern analysis detects 66 specific dystrophin gene restriction
fragments. The multiplex reaction requires 50-fold less DNA than Southern
analysis and thus is considerably more sensitive. SETTING--Fourteen
university-affiliated and private genetic disease diagnostic laboratories.
PATIENTS--Male patients with clinical signs of DMD/BMD. Cases were selected
for analysis randomly, without knowledge of whether a deletion was present
within the dystrophin gene. MAIN OUTCOME MEASURES--The percentage of cases
that were detectable by multiplex PCR in comparison with Southern analysis,
the frequency, extent, and location of the detected deletion mutations. In
some cases, duplication mutations were monitored. RESULTS--The accuracy of
a single PCR multiplex amplification (nine exons) was compared with
Southern analysis with 10 cDNA probes that cover the full length of the
gene. The multiplex PCR analytic method detected 82% of those deletions
detected by Southern analysis methods. In one of 745 analyses, the
multiplex method suggested a single exon deletion, which was not confirmed
by Southern analysis, representing a false-positive rate of 0.013%.
CONCLUSIONS--Multiplex PCR represents a sensitive and accurate method for
deletion detection of 46% of all cases of DMD/BMD. The method requires 1
day for analysis, is easy to perform, and does not use radioactive tracers.
As such, multiplex PCR represents an efficient and rapid method for
prenatal or postnatal diagnosis of DMD/BMD.
