Reactive sequencing of DNA (RSDNA)

Arizona State University-Tempe Campus, Tempe AZ

Investigators

Abstract

DESCRIPTION (provided by applicant): The objective of this project is to develop a DNA sequencing method, which will allow us to sequence thousands of DNA templates simultaneously. This can be done by repeated extension of the primer of a primer-template duplex, which has been covalently bound to a standard glass slide. Known primer sequences are immobilized to the slide in unique "spots" on the surface, which are then allowed to capture and anneal to templates containing a complementary sequence. Sequences can then be read by probing with exonuclease deficient polymerase in the presence of single, fluorescently labeled de-oxy nucleotide tri-phosphates (dNTP's). The spots are subjected to fluorescence imaging to determine which primers have been extended. Before the next reaction can proceed, the fluorescent label must be destroyed. This is done by free radical reaction between the excited fluorescein molecule and diphenyliodonium chloride. Once the fluorescence signal returns to baseline levels, the cycle may begin again.