This Phase I SBIR project investigates a new approach to fluorescence detection for automated DNA sequencing using simple, relatively low cost optics and electronics. Our approach uses reliable, solid state components including diode lasers and is readily scaled from single channel to multichannel (i.e., capillary arrays or multi-lane gels) DNA base calling. We envision commercial instruments based on our technology that will be simpler and more rugged than existing instruments, yet cost less. Automated DNA base calling uses fluorescence lifetime discrimination measured with a novel heterodyne technique. Up to four different dyes can be identified - as can dye mixtures - using only a single detector element and low cost electronics for each elution channel. The Phase I project will demonstrate our approach by identifying and distinguishing DNA primers tagged with recently developed dyes that absorb red light and have lifetimes of l to 5 nsecs. Characterization of detection limits, dynamic range and noise sources as well as determination of the accuracy and precision of fluorescent lifetimes will be performed using a single channel instrument. The Phase II effort will focus on adapting the technique to capillary array systems.
Automated DNA sequencing has application to genetic testing for medical diagnosis, forensics, paternity determination, and a variety of research needs including genome sequencing, mutation analysis, linkage analysis, and gene mapping. If successful, our approach will significantly lower the cost of DNA sequencing instrumentation.
