Efficient synthesis of double dye-labeled oligodeoxyribonucleotide probes and their application in a real time PCR assay
- 15 February 1998
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 26 (4) , 1026-1031
- https://doi.org/10.1093/nar/26.4.1026
Abstract
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 dye-labeled support compared with traditional post-synthetic attachment of rhodamine.Keywords
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