We propose to continue our technology development to develop new instrumentation that will allow extensive automation for a high throughput DNA sequencing facility at low cost. The high throughput at low cost will greatly accelerate the Human Genome Sequencing Project. The design of the fully automated system should fit into 3,000 square feet of laboratory space which further decreases cost and allows more facilities to acquire this system. The goal is to establish such a sequencing system that can accomplish 1 Mb of finished sequence per day at a cost of 1 cent per finished base. The instrumentation includes a point sink DNA shearing device, robotic plaque/colony picker, a universal mobile microtiter plate cassette server, an oxygen enriched microtiter plate cassette shaker, M13 template preparation instrument, plasmid template preparation instrument for sub microliter PCR reactions, an instrument, a coupled reaction pippeting and thermal cycler instrument for sub microliter PCR reactions, an instrument for direct loading into capillary electrophoresis of reaction mixes, a 1,000 capillary electrophoresis instrument with replaceable matrix, a high throughput 96-well plate automated oligonucleotide synthesizer, and automatic finishing instrument, a BAC screening instrument for contig assembly, a micro capillary thermal cycler coupled to a high through-put HPLC instrument for conducting allelic variation, and a BAC mapping instrument for sequence validation. Also contained in this Center is a project for instrumentation, a project for implementation of the equipment for extensive testing, a pilot human genome sequencing project, a project for sequence annotation, validation and allelic variation, a project on informatics and software development for the automated instrumentation, a core facility for high throughput oligonucleotide synthesis, a development proposal is to develop tests and implement a high throughput sequencing facility to generate hundreds of megabases of human DNA sequence per year, and to evaluate all of the recognized coding regions and control regions for allelic variation with a sample of 96 individuals representing the worlds population.
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