|Genome Informatics Section
DOE Human Genome Program Contractor-Grantee Workshop
|110a. Expert System
for Long-Read Base-Calling in DNA Sequencing by Capillary Electrophoresis
Arthur W. Miller and Barry L. Karger
We have recently reported the routine sequencing of 1000 bases in less than one hour by capillary electrophoresis (CE) with replaceable linear polyacrylamide solutions (Salas-Solano et al., Anal. Chem. 1998, 70, 3996-4003). One factor contributing to this result was a base-calling expert system, ABC. Compared to our earlier base-calling approaches, the principal benefit of this base-caller has been a reduction in errors at read lengths above 800 bases, where peaks may be too poorly resolved to determine precise base positions. A more flexible and robust version of ABC has now been developed, which begins by performing color separation and baseline subtraction. It then divides the electropherogram into short sections, which are analyzed independently to estimate noise, peak width, and other parameters. This initial analysis is used to select basecalling rules for each region of the data, which are applied to determine the final DNA sequence. Base confidences are assigned using decision trees. ABC works with four-dye CE or slab gel data acquired using four or more raw spectral channels, and requires no user configuration.
This work is being supported by DOE grant DE-FG02-98ER 69895.
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