Focal Extraction of Surface-Bound DNA from a Microchip Using Photo-Thermal Denaturation

Abstract

High-throughput, selective extraction of a particular DNA fragment from a mixture of DNA before PCR amplification is becoming increasingly important in the DNA analysis field. Although the latest microchip technology has enabled real-time DNA expression analysis using hybridization between surface-bound probe DNA and sample DNA, the potential of this technology in purification of a small amount of DNA has not been demonstrated. We report here a method for area-selective release and collection of specific DNA, in which an IR laser beam is focused onto surface-bound sample DNA at the target-spotted area to denature hybridized DNA. First, sample DNA labeled with a fluorescent dye was hybridized to a probe DNA immobilized on a chromiumcoated chip. A 1053-nm IR laser beam with an intensity of 10–100 mW was then focused on the target area with a spatial resolution of 10μm, causing the release of the fluorophore-labeled sample DNA as a result of photo-thermal denaturation. Confirmation of the amount of eluted DNA by PCR amplification after collection indicated that more than 10-20 mol DNA/μm2 area was eluted from the microchip, representing more than 70% of the chip-bound sample DNA. These results indicate that this method can be applied to the highly sensitive purification of DNA in microchip technology.