This proposal has as its goal the development of capillary gel electrophoresis for the high throughput/high resolution analysis of single stranded DNA fragments for sequencing. The investigators will develop columns and conditions to resolve roughly 500 bases per hour per column, with baseline resolution of species differing by a single base. While they will initially focus on l - 300 base units of deoxyoligonucleotides, they will devote significant efforts to maintain single base resolution up to l000 base units with appropriate capillary columns. Detection will be based on laser induced fluorescence using an argon-ion laser with detection limits at the attomole level. The system will be interfaced to a robotic sample pretreatment instrument that has already been shown to handle the chemistry of sequencing. The applicants will also develop an instrument utilizing multiple columns for a significant increase in analytical throughput. They will demonstrate feasibility on samples of known sequence and blind samples at all development stages.
| Huang, X C; Stuart, S G; Bente 3rd, P F et al. (1992) Capillary gel electrophoresis of single-stranded DNA fragments with UV detection. J Chromatogr 600:289-95 |
