Genotyping by dynamic heating of monolayered beads on a microheated surface

• Published in: Electrophoresis Vol. 25; no. 21-22; pp. 3712 - 3719
• Main Authors: Russom, Aman, Haasl, Sjoerd, Ohlander, Anna, Mayr, Torsten, Brookes, Anthony J., Andersson, Helene, Stemme, Göran
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 01.11.2004; WILEY‐VCH Verlag
• Subjects: Alleles; Beads; Clinical genetics; Dermatologi och venerologi, klinisk genetik, invärtesmedicin; Dermatology and venerology,clinical genetics, internal medicine; DNA Mutational Analysis - methods; Dynamic allele-specific hybridization; Equipment Design; Genotype; Hot Temperature; Klinisk genetik; Lab-On-A-Chip Devices; MEDICIN; Medicin och hälsovetenskap; MEDICINE; Microchip Analytical Procedures - methods; Microcontact printing; Microheater; Microspheres; Miniaturization; Nucleic Acid Denaturation; Nucleic Acid Hybridization; Oligonucleotides - analysis; Oligonucleotides - genetics; Polymorphism, Single Nucleotide; Single-nucleotide polymorphism
• ISSN: 0173-0835; 1522-2683; 1522-2683
• DOI: 10.1002/elps.200406065

A miniaturized bead‐based dynamic allele‐specific hybridization (DASH) approach for single‐nucleotide polymorphism analysis is presented. Chips with integrated heater and temperature sensors for open‐surface DNA analysis were microfabricated. Microcontact printing using a poly(dimethylsiloxane) (PDMS) stamp was employed to create monolayers of immobilized beads on the surface of the chip. This chip allows fast, well‐controllable temperature ramping. The temperature distribution was homogeneous over the entire heater area. All three possible variants of an SNP site of a synthesized oligonucleotide were accurately scored using the bead‐based DASH approach. Our assay has a nonoptimized temperature ramping rate of 4°C–6°C/min compared to earlier reported values of 2°C–3°C/min, thereby reducing the total analysis time by a factor of 2. Reliable DASH measurement data from areas as small as 12×13 μm was achieved. Our bead‐based DASH approach has enabled a dramatic volume reduction and is a step towards developing a cost‐effective high‐throughput DASH method on arrays of single beads.