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Nucleic Acids Research, Vol 25, Issue 22 4500-4504, Copyright © 1997 by Oxford University Press

ARTICLES

New dye-labeled terminators for improved DNA sequencing patterns

BB Rosenblum, LG Lee, SL Spurgeon, SH Khan, SM Menchen, CR Heiner and SM Chen
PE Applied Biosystems, 850 Lincoln Centre Drive, Foster City, CA 94404, USA. rosenbbb@perkin-elmer.com

We have used two new dye sets for automated dye-labeled terminator DNA sequencing. One set consists of four, 4,7-dichlororhodamine dyes (d- rhodamines). The second set consists of energy-transfer dyes that use the 5-carboxy-d-rhodamine dyes as acceptor dyes and the 5- or 6-carboxy isomers of 4'-aminomethylfluorescein as the donor dye. Both dye sets utilize a new linker between the dye and the nucleotide, and both provide more even peak heights in terminator sequencing than the dye- terminators consisting of unsubstituted rhodamine dyes. The unsubstituted rhodamine terminators produced electropherograms in which weak G peaks are observed after A peaks and occasionally C peaks. The number of weak G peaks has been reduced or eliminated with the new dye terminators. The general improvement in peak evenness improves accuracy for the automated base-calling software. The improved signal-to-noise ratio of the energy-transfer dye-labeled terminators combined with more even peak heights results in successful sequencing of high molecular weight DNA templates such as bacterial artificial chromosome DNA.
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