Propanoic acid, 2,2-dimethyl-, 1,1'-[6-[10-[bis(1-methylethyl)amino]-13-cyano-1-oxo-9,11-dioxa-2-aza-10-phosphatridec-1-yl]-2',4,4',5',7,7'-hexachloro-3-oxospiro[isobenzofuran-1(3H),9'-[9H]xanthene]-3',6'-diyl] ester Chemical Properties

Boiling point:

928.7±65.0 °C(Predicted)

storage temp. 

-20°C

solubility 

Good solubility in acetonitrile and DCM

pka

12.74±0.20(Predicted)

color 

white to off-whitepowder

Appearance

off white solid

Propanoic acid, 2,2-dimethyl-, 1,1'-[6-[10-[bis(1-methylethyl)amino]-13-cyano-1-oxo-9,11-dioxa-2-aza-10-phosphatridec-1-yl]-2',4,4',5',7,7'-hexachloro-3-oxospiro[isobenzofuran-1(3H),9'-[9H]xanthene]-3',6'-diyl] ester Usage And Synthesis

Description

HEX phosphoramidite for oligonucleotide synthesis, pure 6-isomer.

HEX (hexachlorofluorescein) is a fluorescein derivative with emission in the yellow spectrum range (absorption maximum at 533 nm, emission maximum at 549 nm).

HEX phosphoramidite is used for synthesis of fluorescent-labeled primers and hybridization probes such as TaqMan, Molecular Beacon, and Scorpion for qPCR. HEX is most effectively quenched by non-fluorescent DusQ 1 dark quencher because of significant overlapping of their spectra (convenient for use with DusQ 1 CPG 500 solid support with a pore size of 500 Å). Many automated sequencers based on capillary gel electrophoresis have a detection channel for HEX. Therefore, this phosphoramidite is commonly used for synthesis of 5’-labeled oligonucleotides for fragment analysis, particularly for microsatellite analysis, when microsatellite loci are amplified using a fluorescent-labeled forward primer and a non-labeled reverse primer.

Coupling: 3 min.

Deprotection: standard conditions using 25% ammonium; deprotection time depends on oligonucleotide composition and nucleobase protecting groups (deprotection for 17 h at 55 °С removes all protecting groups from standard nucleobases). AMA (solution of concentrated aqueous ammonium/40% aqueous methylamine 1:1 v/v) can be used with ~5% of 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 (e.g. 10 min at 65 °C).

Uses

6-Hexachloro-fluorescein phosphoramidite is a fluorescent probe that can be used for oligonucleotide labeling[1].

References

[1] Lee HB, et al. Allele-Specific Quantitative PCR for Accurate, Rapid, and Cost-Effective Genotyping. Hum Gene Ther. 2016 Jun;27(6):425-35. DOI:10.1089/hum.2016.011

Abs/Em Maxima

533/549 nm

Fluoresene quantum yield

0.57

Extinction Coefficient

87770

CF260

0.3

CF280

0.13

Propanoic acid, 2,2-dimethyl-, 1,1'-[6-[10-[bis(1-methylethyl)amino]-13-cyano-1-oxo-9,11-dioxa-2-aza-10-phosphatridec-1-yl]-2',4,4',5',7,7'-hexachloro-3-oxospiro[isobenzofuran-1(3H),9'-[9H]xanthene]-3',6'-diyl] ester(1360547-55-2)Related Product Information

• 6-TET phosphoramidite [5'-Tetrachlorofluorescein phosphoramidite]
• 5'(6)-FAM