title: Identification of deletion and duplication genotypes of the PMP22 gene using PCR-RFLP, competitive multiplex PCR, and multiplex ligation-dependent probe amplification: a comparison.
abstract: 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.
authors.affiliation: Institute of Biomedical Engineering, College of Medicine and College of Engineering, National Taiwan University, Taipei, Taiwan.
authors.forename: Chia-Cheng
authors.initials: CC
authors.lastname: Hung
authors.forename: Chien-Nan
authors.initials: CN
authors.lastname: Lee
authors.forename: Chia-Yun
authors.initials: CY
authors.lastname: Lin
authors.forename: Wen-Fang
authors.initials: WF
authors.lastname: Cheng
authors.forename: Chi-An
authors.initials: CA
authors.lastname: Chen
authors.forename: Sung-Tsang
authors.initials: ST
authors.lastname: Hsieh
authors.forename: Chih-Chao
authors.initials: CC
authors.lastname: Yang
authors.forename: Yuh-Jyh
authors.initials: YJ
authors.lastname: Jong
authors.forename: Yi-Ning
authors.initials: YN
authors.lastname: Su
authors.forename: Win-Li
authors.initials: WL
authors.lastname: Lin
mesh_terms: D016022:Case-Control Studies; D002607:Charcot-Marie-Tooth Disease / Q000235:genetics; D017353:Gene Deletion; D018628:Gene Dosage; D020440:Gene Duplication; D005838:Genotype; D015417:Hereditary Sensory and Motor Neuropathy / Q000235:genetics; D006801:Humans; D015336:Molecular Probe Techniques; D009185:Myelin Proteins / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length
chemical_list: D009185:Myelin Proteins; C073687:PMP22 protein, human