title: Validation of GNE:p.M712T identification by melting curve analysis.
abstract: Hereditary inclusion body myopathy/distal myopathy with rimmed vacuoles is an adult onset autosomal recessive muscle-wasting disease common in people of Iranian-Jewish descent, due to the founder allelic variant GNE:p.M712T. High correlation of disease susceptibility with GNE:p.M712T allows its use as a molecular marker for diagnosis. In this study, we applied and validated the use of melting curve analysis using SimpleProbe technology for detection of this mutation using specimens obtained by mouthwash, buccal swab, and whole blood. The assay was then applied to 43 clinical specimens, and results were validated by additional methods. A probe spanning this mutation in exon 12 accurately discerns two Tm corresponding to its hybridization to wild-type and M712T-derived amplicons. A 10 degrees C divergence in Tm allowed rapid single-tube genotyping of reference and patient samples with 100% accuracy. Distal myopathy constitutes a large heterogeneous group of pathologies with similar physiological manifestations and little molecular markers for distinguishing subtypes. Application of SimpleProbes for detection of GNE:p.M712T on genomic DNA obtained from buccal epithelial cells allows accurate, rapid, and cost-effective identification of this allele in individuals at risk. This procedure is amenable to automated high-throughput applications and can be extended to both clinical and research applications.
authors: HIBM Research Group, Encino, California, USA.
authors: Yadira
authors: Y
authors: Valles-Ayoub
authors: Chai
authors: C
authors: Saechao
authors: Arman
authors: A
authors: Haghighatgoo
authors: Mehran S
authors: MS
authors: Neshat
authors: Saghi
authors: S
authors: Esfandiarifard
authors: Marvin
authors: M
authors: Pietruszka
authors: Daniel
authors: D
authors: Darvish
mesh_terms: D000328:Adult; D000483:Alleles; D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D004252:DNA Mutational Analysis / Q000379:methods*; D049310:Distal Myopathies / Q000201:enzymology / Q000235:genetics; D018703:Founder Effect; D006801:Humans; D015336:Molecular Probe Techniques; D009097:Multienzyme Complexes / Q000235:genetics*; D018979:Myositis, Inclusion Body / Q000201:enzymology / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D013816:Thermodynamics
chemical_list: D009097:Multienzyme Complexes; C482901:UDP-N-acetylglucosamine 2-epimerase - N-acetylmannosamine kinase; D004247:DNA