High-speed DNA sequencing: an approach based upon fluorescence detection of single molecules.

We are developing a laser based technique for the rapid sequencing of large fragments (approximately 40 kb) of DNA based upon the detection of single, fluorescently tagged nucleotides cleaved from a single DNA fragment. We have demonstrated significant progress on several of the important steps of this technique. The projected rate of sequencing is several hundred bases per second which is orders of magnitude faster than existing methods. Once developed, this technology could be utilized by investigators for rapid sequencing of genetic material from virtually any source.

Ray tracing analysis of the image quality of a high collection efficiency mirror system.

Recently, a high collection efficiency mirror system was developed by Watson [Cytometry 10, 681-688 (1989)] to increase the sensitivity of low level fluorescence detection. The mirror system consists of an ellipsoidal imaging mirror and spherical backreflecting mirror. The fluorescing sample is located at one focus of the ellipsoid, and its image is formed at the other focus. In this paper we evaluate the image quality of this geometry using a PC-based ray tracing program. The analysis demonstrates high collection efficiency but poor image quality. The effect of poor image quality on single molecule detection is discussed.