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Published online 23 January 2004

Nucleic Acids Research, 2004, Vol. 32, No. 2 e18
© 2004 Oxford University Press

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

James H. Watterson, Sandeep Raha, Christopher C. Kotoris, Christopher C. Wust, Farhad Gharabaghi, Sarah C. Jantzi, Nicole K. Haynes, Nathalie H. Gendron1, Ulrich J. Krull, Alex E. Mackenzie1 and Paul A. E. Piunno*

Chemical Sensors Group, University of Toronto at Mississauga, 3359 Mississauga Road North, Mississauga, Ontario, Canada L5L 1C6 and 1 Children’s Hospital of Eastern Ontario Research Institute, Molecular Genetics Laboratory, 401 Smyth Road, Ottawa, Ontario, Canada K1H 8L1

*To whom correspondence should be addressed. Tel: +1 905 828 5453; Fax: +1 905 828 5425; Email: ppiunno@utm.utoronto.ca

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 (~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.


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