title: Determination of SMN1/SMN2 gene dosage by a quantitative genotyping platform combining capillary electrophoresis and MALDI-TOF mass spectrometry.
abstract: BACKGROUND
  Spinal muscular atrophy (SMA) is a common inherited and fatal neuromuscular disease caused by deletions and/or mutations that lead to altered concentrations of proteins encoded by the survival motor neuron genes SMN1 and SMN2. Because of the high incidence (at least 1 in 10,000 live births and a carrier frequency of 1 in 35 to 1 in 50) and severity of the disease, precise quantification of SMN1 and SMN2 gene copy numbers is essential for diagnosis and genetic counseling.
  
  
  METHODS
  We developed a genotyping platform combining capillary electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to quantify absolute gene dosage. The absolute gene dosage can be determined by a multiplexed competitive PCR protocol followed by capillary electrophoresis analysis. The relative SMN1/SMN2 ratio can be analyzed by PinPoint assay followed by MALDI-TOF MS analysis.
  
  
  RESULTS
  The complementary assays were evaluated in confirmed cases including 9 affected patients, 33 carriers, and 478 healthy individuals from the general population. We were able to determine all genotypes with different SMN1/SMN2 gene copy number ratios, which unambiguously diagnosed carrier status and the severity of SMA with 100% specificity.
  
  
  CONCLUSIONS
  This quantitative genotyping platform is suitable for detection of SMA. The described approach may serve as a general quantitative genotyping method for molecular diagnosis of other inheritable diseases.
authors.affiliation: Institute of Chemistry, and Genomics Research Center, Academia Sinica, Taipei, Taiwan, ROC.
authors.forename: Hung-Yi
authors.initials: HY
authors.lastname: Kao
authors.forename: Yi-Ning
authors.initials: YN
authors.lastname: Su
authors.forename: Hsin-Kai
authors.initials: HK
authors.lastname: Liao
authors.forename: Ming S
authors.initials: MS
authors.lastname: Liu
authors.forename: Yu-Ju
authors.initials: YJ
authors.lastname: Chen
mesh_terms: D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics*; D019075:Electrophoresis, Capillary; D018628:Gene Dosage; D005820:Genetic Testing; D005838:Genotype; D006579:Heterozygote; D006801:Humans; D009134:Muscular Atrophy, Spinal / Q000175:diagnosis* / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D016601:RNA-Binding Proteins / Q000235:genetics*; D015203:Reproducibility of Results; D055532:SMN Complex Proteins; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014897:Spinal Muscular Atrophies of Childhood / Q000175:diagnosis / Q000235:genetics; D055533:Survival of Motor Neuron 1 Protein; D055540:Survival of Motor Neuron 2 Protein
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D009419:Nerve Tissue Proteins; D016601:RNA-Binding Proteins; D055532:SMN Complex Proteins; C526471:SMN1 protein, human; C526477:SMN2 protein, human; D055533:Survival of Motor Neuron 1 Protein; D055540:Survival of Motor Neuron 2 Protein