One of the major challenges in genome sciences is to identify and understand the role of single nucleotide polymorphisrns (SNPs) in common disease. It is envisioned that new technologies will be developed in providing the necessary tools for accurate diagnosis of inherited disease, the determination risk factors contributing to common disease, and the identification of a patient's metabolic profile. Such technology brings promise in prevention in numerous ways such as prophylactic treatment to delay the onset or progression of disease, and treatment with medication(s) with the safest and most efficacious outcome. Presently however, DNA sequencing technology is not adequate in rapidly identifying SNPs in thousands of individuals, let alone a large portion of our society. Ideally, DNA sequencing technology would interrogate the entire genome sequence directly from a person's genornic DNA in realtime, and analyses would be done in the field. Thus, new technologies in real-time, portable devices are greatly needed for rapid and accurate identification of sequence variation, The second part of interpretation and understanding the role of SNPs is beyond the scope of this proposal, but it is believed that discoveries of specific SNPs in particular genes that represent risk factors or are causative agents of disease will be made in parallel with the proposed research. Here, we propose the construction of a new portable DNA sequencing device with the potential for detection of SNPs directly from genomic DNA sources. To accomplish this, we describe experiments utilizing our PuIse-Multiline Excitation (PME) technology coupled with microfluidic separation chips. The microPME strategy has several advantages over conventional DNA sequencing technology resulting in significantly enhanced fluorescent detection utilizing compact and light-weight laser sources. To test the feasibility of the portable technology, we purpose the construction of a compact PME detector coupled to a multi-channel microfluidic separation chip within the confinement of a suitcase footprint. The instrument will be validated by reconstruction experiments of known SNPs in a multiplex format. Demonstration of feasibility will result in a proposed phase II application to develop an integrated portable DNA sequencer. The successful outcome of these research plans would represent significant increases in throughput and performance, simplified methodologies, and reduced costs. It is anticipated that microPME instruments will have broad applications for routine usage for clinical diagnosis, forensic analyses, military applications, and general sequencing purposes. ? ?
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