Rapid detection of single nucleotide polymorphisms associated with spinal muscular atrophy by use of a reusable fibre-optic biosensor

Nucleic Acids Res. 2004 Jan 23;32(2):e18. doi: 10.1093/nar/gnh013.

Abstract

Rapid (<2 min) and quantitative genotyping for single nucleotide polymorphisms (SNPs) associated with spinal muscular atrophy was done using a reusable (approximately 80 cycles of application) fibre-optic biosensor over a clinically relevant range (0-4 gene copies). Sensors were functionalized with oligonucleotide probes immobilized at high density (approximately 7 pmol/cm2) to impart enhanced selectivity for SNP discrimination and used in a total internal reflection fluorescence detection motif to detect 202 bp PCR amplicons from patient samples. Real-time detection may be done over a range of ionic strength conditions (0.1-1.0 M) without stringency rinsing to remove non-selectively bound materials and without loss of selectivity, permitting a means for facile sample preparation. By using the time-derivative of fluorescence intensity as the analytical parameter, linearity of response may be maintained while allowing for significant reductions in analysis time (10-100-fold), permitting for the completion of measurements in under 1 min.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biosensing Techniques / instrumentation
  • Biosensing Techniques / methods*
  • Cell Line
  • Cyclic AMP Response Element-Binding Protein
  • DNA / analysis
  • DNA / genetics
  • Fiber Optic Technology
  • Genotype
  • Kinetics
  • Muscular Atrophy, Spinal / genetics*
  • Muscular Atrophy, Spinal / pathology
  • Nerve Tissue Proteins / genetics
  • Oligonucleotide Probes / genetics
  • Polymerase Chain Reaction / methods
  • Polymorphism, Single Nucleotide / genetics*
  • RNA-Binding Proteins
  • SMN Complex Proteins
  • Sensitivity and Specificity
  • Time Factors

Substances

  • Cyclic AMP Response Element-Binding Protein
  • Nerve Tissue Proteins
  • Oligonucleotide Probes
  • RNA-Binding Proteins
  • SMN Complex Proteins
  • DNA