AUTOMATED HIGH SPEED DNA SEPARATIONS

DESCRIPTION: Development of high throughput DNA sampling, separation, and detection strategies are described for use in sequencing and genotyping. New technology is proposed for massively parallel, low-volume DNA separations. This will be accomplished by combining capillary sample introduction parallel separations in microfabricated chips, and electrochemical array detection to provide a single integrated system for separation and detection. Ultimately, this will be connected via a sampling capillary to an array of nanovials for introduction of large numbers of samples. The proposal is divided into four major parts which include 1) optimization of separations of DNA fragments in ultrathin channels with sieving buffers, 2) development of an improved interface to allow efficient sample transfer across the capillary-channel interface, 3) development of electrochemical array detection using Ru(bpy)32+ as an electrophore for dsDNA and ssDNA, and 4) application of the capillary transfer method and electrochemical array detection to DNA separations on chips. This will provide the means to rapidly place multiple samples in a very large number of channels (1000 parallel channels is feasible) on a single chip. The proposed technology promises to increase sample throughput by 10- to 50-fold and reduce sample size by 2 to 3 orders of magnitude relative to currently available systems.