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Mullah, Bashar; Livak, Ken; Andrus, Alex; Kenney, Paul
Nucleic acids research, 02/1998, Volume: 26, Issue: 4Journal Article
A fast cleaving non-nucleosidic tetramethylrhodamine dye-labeled support has been developed for automated synthesis of double dye-labeled oligodeoxyribonucleotides in high yield. A mixture (1:1:2) of t-butylamine: methanol:water is used for cleavage and deprotection of dye-labeled oligodeoxyribonucleotides without any degradation or modification of dyes and nucleobases. The cleavage rate of oligodeoxyribonucleotides is significantly increased by using a diglycolate ester linkage instead of the commonly used succinate linkage. These double dye-labeled probes are used in PCR for real time detection of a specific PCR product. Using a 5′-exonuclease assay, detected on the ABI PRISM 7700 Sequence Detection System, there was no distinguishable difference in performance of probes synthesized using the dyelabeled support compared with traditional post-synthetic attachment of rhodamine.
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