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Oligo Synthesis

Oligo Synthesis : CEPs

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Cyanine 5.5 Phosphoramidite

Cyanine 5.5™ Phosphoramidite

Glen Research

Catalogue No.DescriptionPack SizePriceQty
10-5916-02Cyanine 5.5 Phosphoramidite 0.25g £740.00£680.80Offer until : 29-Feb-2020Offer Code : GLEN88% off all Glen Research products View Offer Quantity Add to Order
10-5916-90Cyanine 5.5 Phosphoramidite 100µmoles £360.00£331.20Offer until : 29-Feb-2020Offer Code : GLEN88% off all Glen Research products View Offer Quantity Add to Order
10-5916-95Cyanine 5.5 Phosphoramidite 50µmoles £196.00£180.32Offer until : 29-Feb-2020Offer Code : GLEN88% off all Glen Research products View Offer Quantity Add to Order

Cyanine 5.5 Phosphoramidite

Cyanine 5.5™ Phosphoramidite

Glen Research

Cyanine 5.5™ Phosphoramidite
 

Catalog Number: 10-5916-xx

Description: Cyanine 5.5™ Phosphoramidite

1-[3-(4-monomethoxytrityloxy)propyl]-1'-[3-[(2-cyanoethyl)-
(N,N-diisopropyl)phosphoramidityl]propyl]-3,3,3',3'-tetramethyl-
4,5-benzindodicarbocyanine iodide
Formula: [C68H76N4O4P] I- M.W.: 1171.25 F.W.: 633.74

Diluent: Anhydrous Acetonitrile
Coupling: 3 minute coupling time recommended. Monomers and Cap A Mix that allow for UltraMILD deprotection are preferred. (Catalog Numbers: dA: 10-1601-xx, dC: 10-1015-xx, dG: 10-1621-xx, dT: 10-1030-xx, Cap A: 40-4210-xx/40-4212-xx.) Use 0.02M iodine to avoid degradation of the cyanine dye.
Deprotection: If UltraMILD reagents were used, deprotect at room temperature in 0.05M Potassium Carbonate in Methanol for 4 hrs or Ammonium Hydroxide for 2 hrs. If standard bases were used, deprotection in Ammonium Hydroxide at room temperature for 24-36 hrs will give acceptable yields. However, the oligo will require a reverse phase purification, e.g., Poly Pak. Technical Bulletin
Storage: Freezer storage, -10 to -30°C, dry
Stability in Solution: 2-3 days
 

CYANINE LABELLING

Two cyanine derivatives, Cyanine 3 and Cyanine 5, which differ in structure simply by the number of carbons in the conjugated poly-ene linkage, are joined by the closely related analogues, Cyanine 3.5 and Cyanine 5.5, and are available as phosphoramidites. Cyanine dyes are normally added once at the 5’-terminus and the MMT group should be removed on the synthesizer. The absorbance of the MMT cation (yellow) is noticeably different from the DMT cation (orange), and so, absorbance-based trityl monitors will detect it incorrectly as a low coupling. On the other hand, conductivity detectors will interpret the release more correctly. Cyanine dyes have also been used successfully adjacent to the 3’-terminus as long as 0.02M iodine oxidizer is used during the remainder of the synthesis.

Deprotection of oligos containing Cyanine dyes may be carried out with ammonium hydroxide at room temperature, regardless of the base protecting groups on the monomers used. If there is a need to use ammonium hydroxide at elevated temperature, Cyanine 3 and Cyanine 3.5 are more stable than Cyanine 5 and Cyanine 5.5. However, it is always prudent to use monomers with base labile protecting groups to limit the exposure time to 2 hours or less at 55°C.

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Cyanine 5.5 Phosphoramidite

Cyanine 5.5™ Phosphoramidite

Glen Research

Material Safety Data Sheet

Glen Report 21.2: Deprotection - Volume 3 - Dye-Containing Oligonucleotides

 

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Cyanine 5.5 Phosphoramidite

Cyanine 5.5™ Phosphoramidite

Glen Research

Literature Highlights

Glen Report 21.2: Deprotection - Volume 3 - Dye-Containing Oligonucleotides

Frequently Asked Technical Question

QUESTION: What are the relative extinction coefficients of 5'-Fluorescein, Hex and Tet etc.. at 260 nm and their Lambda max?

RESPONSE:Please see http://www.glenresearch.com/Technical/Extinctions.html

REFERENCE(S):
Oligonucleotide Properties Calculator; http://www.basic.northwestern.edu/biotools/oligocalc.html


QUESTION: What are the relative extinction coefficients of various dyes?

RESPONSE:Please see http://www.glenres.com/Technical/Extinctions.html#dyes


QUESTION: Does AMA or methylamine cause any degradation to fluorescein or fluorescein-type dyes such as FAM or FITC?

RESPONSE:Response: While AMA (Ammonium hydroxide/40% Methylamine 1:1 v/v) is considered compatible with fluorescein, the use of methylamine when deprotecting a Fluorescein-labeled oligo does lead to a small amount of degradation, which is characterized by a the appearance of a late-eluting peak by RP HPLC that shows no visible fluorescein absorbance. With standard deprotection conditions (AMA 10 minutes at 65 C) the amount of degradation is approximately 5%. The impurity is not detected with AMA at RT for 2 hours.

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Cyanine 5.5 Phosphoramidite

Cyanine 5.5™ Phosphoramidite

Glen Research

is not detected with AMA at RT for 2 hours.


DILUTION/COUPLING DATA

The table below shows pack size data and, for solutions, dilutions and approximate couplings based on normal priming procedures. Please link for more detailed usage information with the various synthesizers.

ABI 392/394
Cat.No. Pack
Size
Grams/
Pack
0.1M Dil.
(mL)
LV40 LV200 40nm 0.2µm 1µm 10µm
Approximate Number of Additions
10-5916-02 0.25grams .25grams 2.13 57.67 34.6 21.63 15.73 11.53 2.88
10-5916-90 100µmoles .117grams 1 20 12 7.5 5.45 4 1
10-5916-95 50µmoles .059grams .5 3.33 2 1.25 .91 .67 .17
Expedite
Cat.No. Pack
Size
Grams/
Pack
Dilution
(mL)
Molarity 50nm 0.2µm 1µm 15µm
Approximate Number of Additions
10-5916-02 0.25grams .25grams 3.19 .07 57.4 35.88 26.09 3.59
10-5916-90 100µmoles .117grams 1.5 .07 23.6 14.75 10.73 1.48
10-5916-95 50µmoles .059grams .75 .07 8.6 5.38 3.91 .54
Beckman
Cat.No. Pack
Size
Grams/
Pack
Dilution
(mL)
Molarity 30nm 200nm 1000nm

Approximate Number of Additions
10-5916-02 0.25grams .25grams 3.19 .07 59 36.88 26.82

10-5916-90 100µmoles .117grams 1.5 .07 25.2 15.75 11.45

10-5916-95 50µmoles .059grams .75 .07 10.2 6.38 4.64

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