NEW DYE-LABELED TERMINATORS FOR IMPROVED DNA-SEQUENCING PATTERNS

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-carbox y isomers of 4'-aminomethylfluorescein as the donor dye. Both dye sets utilize a new linker between the dye and the nucleotide, and both pro vide more even peak heights in terminator sequencing than the dye-term inators 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 we ak G peaks has been reduced or eliminated with the new dye terminators . The general improvement in peak evenness improves accuracy for the a utomated base-calling software, The improved signal-to-noise ratio of the energy-transfer dye-labeled terminators combined with more even pe ak heights results in successful sequencing of high molecular weight D NA templates such as bacterial artificial chromosome DNA.