A rapid and reliable detection system for the analysis of PMP22 gene dosage by MP/DHPLC assay

Lin, Chia-Yun; Su, Yi‐Ning; Lee, Chien‐Nan; Hung, Chia-Cheng; Cheng, Wen‐Fang; Lin, Win-Li; Chen, Chi-An; Hsieh, Sung‐Tsang

doi:10.1007/s10038-005-0350-9

Cited by 8 publications

(2 citation statements)

References 33 publications

(34 reference statements)

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“…A wide range of techniques is available for molecular testing of CMT1A/HNPP [18,22,23,[30][31][32][33]. However the faster, cost-effective and reliable method is the analysis of a panel of 15 STRs based on the detection of, at least, two STRs with a trisomic triallelic pattern indicating with certainty the presence of the CMT1A duplication.…”

Section: Discussionmentioning

confidence: 99%

Quantitative Fluorescence-Polymerase Chain Reaction Assay for the Detection of the Duplication of the Charcot Marie Tooth Disease Type 1A Critical Region

Toffol¹,

Bellone

Dulcetti³

et al. 2010

Genetic Testing and Molecular Biomarkers

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Charcot Marie Tooth (CMT) syndrome is the most common hereditary peripheral neuropathy, with an incidence of about 1 in 2500. The subtype 1A (CMT1A) is caused by a tandem duplication of a 1.5-Mb region encompassing the PMP22 gene. Conventional short tandem repeat (STR) analysis can reveal this imbalance if a triallelic pattern, defining with certainty the presence of duplication, is present. In case of duplication with a biallelic pattern, it can only indicate a semiquantitative dosage of the fluorescence intensity ratio of the two fragments. In this study we developed a quantitative fluorescence-PCR using seven highly informative STRs within the CMT1A critical region that successfully disclosed or excluded the presence of the pathogenic imbalance in a cohort of 60 samples including 40 DNAs from samples with the CMT1A duplication previously characterized with two different molecular approaches, and 20 diagnostic samples from 10 members of a five-generation pedigree segregating CMT1A, 8 unrelated cases and 2 prenatal samples. The application of the quantitative fluorescence-PCR using STRs located in the critical region could be a reliable method to evaluate the presence of the PMP22 duplication for the diagnosis and classification of hereditary neuropathies in asymptomatic subjects with a family history of inherited neuropathy, in prenatal samples in cases with one affected parent, and in unrelated patients with a sporadic demyelinating neuropathy with clinical features resembling CMT (i.e., pes cavus with hammer toes) or with conduction velocities in the range of CMT1A.

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Section: Discussionmentioning

confidence: 99%

Quantitative Fluorescence-Polymerase Chain Reaction Assay for the Detection of the Duplication of the Charcot Marie Tooth Disease Type 1A Critical Region

Toffol¹,

Bellone

Dulcetti³

et al. 2010

Genetic Testing and Molecular Biomarkers

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“…Variations in the quality of the extracted DNA can have major effects on amplification efficiency, with the largest effects seen in the larger products in a multiplex reaction. However, our previous multiplex quantitative genotype analysis studies with HPLC instruments showed that competitive PCR allowed the accurate determination of allele dose variations (32,33,50,51 ).…”

Section: Discussionmentioning

confidence: 99%

Quantitative Assay of Deletion or Duplication Genotype by Capillary Electrophoresis System: Application in Prader–Willi Syndrome and Duchenne Muscular Dystrophy

et al. 2006

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Use of multiplex PCR and CE for gene dosage quantification and its biomedical applications for SMN, PMP22, and α‐globin genes

et al. 2007

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Many genetic diseases are caused by the presence of point mutations, small insertions, and deletions in respective genes, and the number of diseases known to be caused by deletions and duplications involving large DNA genomes is increasing. These changes lead to underexpression or overexpression of the gene, according to changes in gene dosage. The methods for the detection of point mutations, small insertions, and deletions are well established, but the detection of larger genomic deletions or duplications is more difficult. Due to the lack of efficient and technically feasible protocols for gene dosage quantification, we describe a diagnostic protocol employing a combination of available methods. The efficient and accurate gene dosage quantification platform is combined with multiplex PCR and CE, and applied to detect dosages of several genes, including SMN, PMP22, and alpha-globin genes. The reliability of this novel methodology shows that it is a relatively speedy and low-cost procedure and a significant tool for genetic diagnosis. Its sensitivity and specificity for identifying deletion and duplication genotypes approach 100%. Moreover, once we establish this powerful system, we will further apply this technique to the rapid detection of trisomy syndromes and microdeletion syndromes, including trisomy 13, Down syndrome, DiGeorge syndrome, and others.

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A rapid and reliable detection system for the analysis of PMP22 gene dosage by MP/DHPLC assay

Cited by 8 publications

References 33 publications

Quantitative Fluorescence-Polymerase Chain Reaction Assay for the Detection of the Duplication of the Charcot Marie Tooth Disease Type 1A Critical Region

Quantitative Fluorescence-Polymerase Chain Reaction Assay for the Detection of the Duplication of the Charcot Marie Tooth Disease Type 1A Critical Region

Quantitative Assay of Deletion or Duplication Genotype by Capillary Electrophoresis System: Application in Prader–Willi Syndrome and Duchenne Muscular Dystrophy

Use of multiplex PCR and CE for gene dosage quantification and its biomedical applications for SMN, PMP22, and α‐globin genes

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