6-TET phosphoramidite [5'-Tetrachlorofluorescein phosphoramidite] Chemical Properties

Boiling point:

895.1±65.0 °C(Predicted)

solubility 

Good solubility in acetonitrile and DCM

pka

12.78±0.20(Predicted)

Appearance

white solid foam

6-TET phosphoramidite [5'-Tetrachlorofluorescein phosphoramidite] Usage And Synthesis

Description

TET phosphoramidite for synthesis of fluorescently labeled oligonucleotides, pure 6-isomer.

TET (tetrachlorofluorescein) is a green-fluorescent fluorescein derivate (absorption maximum at 519 nm, emission maximum at 535 nm).

TET phosphoramidite is used for synthesis of fluorescently-labeled primers and hybridization probes for qPCR. TET can be used with DusQ1 fluorescence quencher (can be used with 500 Å DusQ1 CPG 500).

5’-labeled primers are used with non-labeled reverse primers for microsatellite amplification via PCR followed by fragment analysis. TET-labeled amplification products can be analyzed using various sequencers for capillary electrophoresis, including ABI PRISM^® 310 Genetic Analyzer.

Condensation: 3 min.

Deprotection: standard conditions with 25% ammonium hydroxide; deprotection time depends on oligonucleotide composition and nucleobase protecting groups (deprotection for 17 hours at 55°C removes all protecting groups from standard nucleobases). AMA (solution of 30% ammonium hydroxide/40% aqueous methylamine 1:1 v/v) can be used with ~5% non-fluorescent side product forming. To avoid formation of the side product, start deprotection with ammonium hydroxide (30 min at room temperature), then add an equal volume of 40% aqueous methylamine and continue deprotection as required with AMA (10 min at 65°C).

Uses

6-TET phosphoramidite (5′-Tetrachlorofluorescein phosphoramidite) is a fluorescent dye that can be used for labeling an oligonucleotide with fluorescein[1].

References

[1] El-Hajj HH, et al. Detection of rifampin resistance in Mycobacterium tuberculosis in a single tube with molecular beacons. J Clin Microbiol. 2001 Nov;39(11):4131-7. DOI:10.1128/JCM.39.11.4131-4137.2001

Abs/Em Maxima

519/535 nm

Fluoresene quantum yield

0.47

Extinction Coefficient

100000

CF260

0.17

CF280

0.09

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