Identification of deletion and duplication genotypes of the PMP22 gene using PCR

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Electrophoresis. 2008 Jan 17

Identification of deletion and duplication genotypes of the PMP22

gene using PCR-RFLP, competitive multiplex PCR, and multiplex

ligation-dependent probe amplification: A comparison.

Hung CC, Lee CN, Lin CY, Cheng WF, Chen CA, Hsieh ST, Yang CC, Jong

YJ, Su YN, Lin WL.

Institute of Biomedical Engineering, College of Medicine and College

of Engineering, National Taiwan University, Taipei, Taiwan.

We evaluated the efficacy of PCR-RFLP, competitive multiplex PCR,

and a commercially available system of multiplex ligation-dependent

probe amplification (MLPA) for the determination of deletion and

duplication genotypes of the PMP22 gene. We compared the methods for

efficiency, sensitivity, and specificity. We determined the gene

dosage of the PMP22 gene via PCR-RFLP, competitive multiplex PCR,

and MLPA.

To demonstrate the sensitivity and accuracy of these three methods,

a total of 185 samples from 42 patients with hereditary neuropathy

with liability to pressure palsies (HNPP), 57 patients with Charcot-

Marie-Tooth disease type 1A (CMT1A), and 86 unaffected individuals,

were analyzed.

Molecular diagnosis by PCR-RFLP was performed on all 185 samples; 24

HNPP deletions and 33 CMT1A duplications were identified. In

contrast, 25 HNPP deletions and 38 CMT1A duplications were

identified correctly using competitive multiplex PCR and MLPA. Six

samples were incorrectly identified by PCR-RFLP (one HNPP deletion

and five CMT1A duplications).

Competitive multiplex PCR and MLPA demonstrated reliability and

relative speed compared to PCR-RFLP; they were superior to PCR-RFLP

for gene dosage quantification. Multiplex PCR and MLPA should be the

methods of choice for detection of deletion and duplication

genotypes in molecular genetic diagnoses.