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Oligo Synthesis

Oligo Synthesis : CEPs

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Biotin Phosphoramidite

Biotin Phosphoramidite

Glen Research

Catalogue No.DescriptionPack SizePriceQty
10-1953-02Biotin Phosphoramidite 0.25g £540.00£496.80Offer until : 29-Feb-2020Offer Code : GLEN88% off all Glen Research products View Offer Quantity Add to Order
10-1953-90Biotin Phosphoramidite 100µmoles £236.00£217.12Offer until : 29-Feb-2020Offer Code : GLEN88% off all Glen Research products View Offer Quantity Add to Order
10-1953-95Biotin Phosphoramidite 50µmoles £132.00£121.44Offer until : 29-Feb-2020Offer Code : GLEN88% off all Glen Research products View Offer Quantity Add to Order

Biotin Phosphoramidite

Biotin Phosphoramidite

Glen Research

Biotin Phosphoramidite

Structure

Catalog Number: 10-1953-xx

Description: Biotin Phosphoramidite

1-Dimethoxytrityloxy-2-(N-biotinyl-4-aminobutyl)-propyl-3-O-
(2-cyanoethyl)-(N,N-diisopropyl)-phosphoramidite
Formula: C47H66N5O7PS M.W.: 876.10 F.W.: 435.48

Diluent: Anhydrous Acetonitrile
Coupling: 15 minute coupling time recommended
Deprotection: No changes needed from standard method recommended by synthesizer manufacturer.
Storage: Refrigerated storage, maximum of 2-8°C, dry

Stability in Solution: 2-3 days

BIOTIN LABELLING

Glen Research biotin phosphoramidites for direct labelling of synthetic oligonucleotides exhibit the following features:

1. All are soluble in acetonitrile at concentrations useful for DNA synthesis.

2. All include a DMT group for cartridge purifications which is essential for the preparation of biotinylated PCR primers because of the potential for cross contamination in HPLC purifications.

3. For the development of diagnostic probes, biotin phosphoramidite is capable of branching to allow multiple biotins to be introduced at the 3’- or 5’-terminus while biotin-dT can replace dT residues within the oligonucleotide sequence. BiotinTEG Phosphoramidite contains a 15 atom mixed polarity spacer arm based on a triethylene glycol. 5’-Biotin phosphoramidite can be added ONLY ONCE to the 5’-terminus of an oligonucleotide. However, the DMT group on the biotin can be used in RP cartridge and HPLC purification techniques.

4. Protected Biotin Serinol Phosphoramidite and CPG are protected with a t-butylbenzoyl group on the biotin ring. This group is designed to stop any phosphoramidite reactions at this active position in biotin. This protection avoids branching when using nucleophilic activators like DCI. The protecting group is easily removed during oligonucleotide cleavage and deprotection. The BiotinLC versions are similarly protected and should be useful for the synthesis of highly sensitive biotinylated probes.

 

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Biotin Phosphoramidite

Biotin Phosphoramidite

Glen Research

Material Safety Data Sheet

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Biotin Phosphoramidite

Biotin Phosphoramidite

Glen Research

REFERENCE(S):
(1) M.L. Fontanel, H. Bazin, and R. Teoule, Analytical Biochemistry, 1993, 214, 338-340.

If you cannot find the answer to your problem below then please contact us or telephone 01954 210 200

Biotin Phosphoramidite

Biotin Phosphoramidite

Glen Research

Frequently Asked Technical Question

QUESTION: Can oligonucleotides modified at the 5'-terminus with, for example, biotin be phosphorylated with kinase?

RESPONSE:Modification reagents based on a 1,2-diol, e.g., BioTEG, DNP phosphoramidites, or a 1,3-diol, e.g., fluorescein, biotin phosphoramidites, can be added several times at the 5'-terminus since they contain an alcohol group capable of further addition with phosphoramidites. Can this alcohol also be as a substrate for T4 polynucleotide kinase for 32P labelling of these modified oligonucleotides? Surprisingly, the answer is yes. Teoule and coworkers have shown(1) that oligos labelled at the 5'-terminus are substrates for kinase. Interestingly, the oligos modified with reagents based on 1,2-diols are labelled to 50%, indicating that only one diastereomer is labelled, while those modified with 1,3-diol reagents are labelled to 100%.

REFERENCE(S):
(1) M.L. Fontanel, H. Bazin, and R. Teoule, Analytical Biochemistry, 1993, 214, 338-340.


QUESTION: What absorbance does biotin have at 260nm? The HPLC trace shows absorbance at 254nm?

RESPONSE:Biotin is transparent at 260nm. The UV detector in the HPLC trace of biotin phosphoramidites is monitoring the absorption of the DMT group on the spacer arm.


QUESTION: Do you have a biotin phosphoramidite containing a disulfide linker which can be cleaved later with DTT to release the DNA from a streptavidin support?

RESPONSE:No. However, this can be produced on the synthesizer by adding to the 5'- terminus first 5'-thiol-modifier C6 S-S (10-1936) followed by BioTEG phosphoramidite (10-1955). This should generate a biotinylated primer with a long spacer arm containing the disulfide linkage which can be cleaved later with DTT.


QUESTION: How can I tell if the biotinylated oligonucleotide I have made really does contain biotin?

RESPONSE:A colorimetric assay for biotin can be quite effective. The color results from the reaction of biotin with p-dimethylaminocinnamaldehyde in the presence of sulfuric acid.

1. Spot 0.2 A260 units of biotinylated oligonucleotide on a silica gel TLC plate.

2. Dry the plate.]

3. Spray with a solution of 2% p-dimethylaminocinnamaldehyde (Sigma), 2% conc. sulfuric acid in ethanol.

4. Heat the plate and the presence of biotin will be indicated by the formation of a pink spot.

Since the intensity of the biotin spot is quite low, it is prudent to compare with an unlabelled oligonucleotide similarly treated.


QUESTION: Can oligonucleotides modified at the 5'-terminus with, for example, biotin be phosphorylated with kinase?

RESPONSE:Modification reagents based on a 1,2-diol, e.g., BioTEG, DNP phosphoramidites, or a 1,3-diol, e.g., fluorescein, biotin phosphoramidites, can be added several times at the 5'-terminus since they contain an alcohol group capable of further addition with phosphoramidites. Can this alcohol also be as a substrate for T4 polynucleotide kinase for 32P labelling of these modified oligonucleotides? Surprisingly, the answer is yes. Teoule and coworkers have shown(1) that oligos labelled at the 5'-terminus are substrates for kinase. Interestingly, the oligos modified with reagents based on 1,2-diols are labelled to 50%, indicating that only one diastereomer is labelled, while those modified with 1,3-diol reagents are labelled to 100%.

REFERENCE(S):
(1) M.L. Fontanel, H. Bazin, and R. Teoule, Analytical Biochemistry, 1993, 214, 338-340.

If you cannot find the answer to your problem below then please contact us or telephone 01954 210 200

Biotin Phosphoramidite

Biotin Phosphoramidite

Glen Research

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
10-1953-02 0.25grams .25grams 2.85 81.67 49 30.63 22.27 16.33 4.08
10-1953-90 100µmoles .088grams 1 20 12 7.5 5.45 4 1
10-1953-95 50µmoles .044grams .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
10-1953-02 0.25grams .25grams 4.26 .07 78.8 49.25 35.82 4.93
10-1953-90 100µmoles .088grams 1.5 .07 23.6 14.75 10.73 1.48
10-1953-95 50µmoles .044grams .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
10-1953-02 0.25grams .25grams 4.26 .07 80.4 50.25 36.55

10-1953-90 100µmoles .088grams 1.5 .07 25.2 15.75 11.45

10-1953-95 50µmoles .044grams .75 .07 10.2 6.38 4.64

If you cannot find the answer to your problem below then please contact us or telephone 01954 210 200