Dissection and Acquisition of DNA Fragments Based on Microsystems

Abstract

A method based on microsystems for the physical dissection of DNA and the recovery of the dissected fragment is developed for the acquisition of an aimed position on a DNA strand. The device consists of a glass substrate, onto which a sacrificial layer, DNA carrier layer, and a pair of electrodes are deposited. Fluorescence labeled DNA is electrostatically stretched and immobilized onto the carrier layer with one of its molecular end aligned on the electrode edge. Using an AFM probe as a knife, an aimed portion of the DNA together with the carrier layer is dissected under the fluorescence microscope. By dissolving the sacrificial layer, the DNA fragment on the carrier piece is recovered onto a membrane filter. The carrier layer is then melted to obtain DNA fragments in solution. The yield of DNA recovery in such a device is investigated by PCR assay, and fluorescence labeling is identified as a major factor that may deteriorate the yield, in particular when DNA is dried and irradiated by the excitation light. A new DNA labeling is developed, where the fluorescence dye is incorporated only to the molecular ends, and the yield close to unity is demonstrated.