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Science 16 October 1987:
Vol. 238. no. 4825, pp. 336 - 341
DOI: 10.1126/science.2443975
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Articles

Science, Vol 238, Issue 4825, 336-341
Copyright © 1987 by American Association for the Advancement of Science

articles

A system for rapid DNA sequencing with fluorescent chain-terminating dideoxynucleotides

JM Prober, GL Trainor, RJ Dam, FW Hobbs, CW Robertson, RJ Zagursky, AJ Cocuzza, MA Jensen, and K Baumeister

Engineering Physics Laboratory, E.I. du Pont de Nemours & Company (Inc.), Wilmington, DE 19898.

A DNA sequencing system based on the use of a novel set of four chain-terminating dideoxynucleotides, each carrying a different chemically tuned succinylfluorescein dye distinguished by its fluorescent emission is described. Avian myeloblastosis virus reverse transcriptase is used in a modified dideoxy DNA sequencing protocol to produce a complete set of fluorescence-tagged fragments in one reaction mixture. These DNA fragments are resolved by polyacrylamide gel electrophoresis in one sequencing lane and are identified by a fluorescence detection system specifically matched to the emission characteristics of this dye set. A scanning system allows multiple samples to be run simultaneously and computer-based automatic base sequence identifications to be made. The sequence analysis of M13 phage DNA made with this system is described.


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