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TERMINUS MODIFIERSGlen Research 5'-Modifiers are designed for use in DNA synthesizers to functionalize the 5'-terminus of the target oligonucleotide. The 5'-Amino-Modifiers are available with a variety of chain lengths to fit exactly the desired application. The DMS(O)MT-protected amino group is easier to deprotect compared to the MMT-protected one. The sulfoxy derivative survives conditions of oligonucleotide synthesis and can either be cleaved with standard deblock solution, or left intact for HPLC purification. At the same time, the DMS(O)MT group is fully compatible with cartridge purification. When detritylation on a cartridge is carried out, the DMS(O)MT+, which is more stable than MMT+, does not reattach itself to an amine. We now offer 5'-DMS(O)MT-Amino-Modifier C6 utilizing this new trityl based protecting group. 5'-Amino-Modifier TEG, a hydrophilic triethylene glycol ethylamine derivative, is 12 atoms in length and fully soluble in aqueous media.
Our more recent 5'-amino modifiers, protected by a novel phthalic acid diamide (PDA) protecting group, are stable solids . In contrast to the TFA protected amino modifiers, which are viscous oils, the analogous PDA protected compounds are granular powders. This important property of these compounds allows straightforward handling, storage and aliquoting and leads to a significant increase in stability. Deprotection with methylamine in gas phase or aqueous solution or AMA leads to fast and complete removal of the PDA protecting group. However, ammonium hydroxide will not drive the equilibrium reaction to completion and only partial deprotection occurs - overnight deprotection with ammonium hydroxide will yield around 80% active amine. We are offering three PDA Amino-Modifiers:
The disulfide thiol modifier may be used for introducing 3'- or 5'-thiol linkages. Dithiol Phosphoramidite (DTPA) is a disulfide-containing modifier designed to functionalize synthetic DNA or RNA with multiple thiol groups and can be incorporated at any position of the oligonucleotide. Each DTPA addition leads to two thiol groups. This modifier was designed for optimal tethering of oligonucleotides to a gold surface but it can also be used for multiple reactions with maleimides and other thiol-specific derivatives. 5'-Carboxy-Modifier C10 is a unique linker designed to be added at the terminus of an oligonucleotide synthesis. It generates an activated carboxylic acid N-hydroxysuccinimide (NHS) ester suitable for immediate conjugation on the synthesis column with molecules containing a primary amine, resulting in a stable amide linkage. PC Amino-Modifier is a photocleavable C6 amino-modifier, part of our line of photocleavable (PC) modifiers. 5'-AminoOxy-Modifier 11 is based on a tetraethylene glycol linkage for improved solubility and for reducing the potential negative impact on hybridization of the oligo. The oxime formed from the reaction of alkyloxyamines with aldehydes creates a stable covalent bond. In comparison, the imine formed by the conjugation of primary amines with aldehydes is not stable to acidic or basic conditions and requires subsequent reduction with borohydride to form stable amine conjugates. 5'-Maleimide Modifier Phosphoramidite, developed at the University of Barcelona, incorporates a maleimide cycloadduct that is stable to ammonium hydroxide at room temperature. This phosphoramidite can be incorporated into DNA and RNA with both phosphate and phosphorothioate linkages. A retro–Diels-Alder reaction deprotects the maleimide immediately prior to conjugation.
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INTELLECTUAL PROPERTYDTPA and derived products (Chemical Products) are for research purposes only, and may not be used for commercial, clinical, diagnostic or any other use. Commercial oligo houses are constrained to sell oligos containing DTPA for research purposes only. These products or portions thereof are subject to proprietary rights of FRIZ Biochem Gesellschaft für Bioanalytik mbH. US Patent No. 7,601,848 and European Patent Application No. 02799716.2. 5'-Maleimide Modifier Phosphoramidite is protected by a patent application and is offered by Glen Research under a non-exclusive license agreement from the University of Barcelona. SEE ALSOPC modifiers p72
ABBREVIATIONSCNEt = Cyanoethyl OTHER INSTRUMENT TYPESAll minor bases, RNA products and modifiers are packaged in septum-capped vials suitable for ABI and other instruments. If you would like another type of vial/column add the following to the end of the catalog number.
SEE ALSOSEE ALSO |
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SEQUENCE MODIFIERSSequence Modifiers are designed for use in automated synthesis.
The carboxy-dT is hydrolyzed during deprotection and can be coupled directly
to a molecule containing a primary amino group by a standard peptide coupling
or via the intermediate N-hydroxysuccinimide (NHS) ester. Amino-Modifier
dA, Amino-Modifier dC, Amino-Modifier dG and both Amino-Modifier dT products
can be added in place of a dA, dC, dG and dT residue, respectively, during
oligonucleotide synthesis. Corresponding Amino-Modifier supports can replace
their respective deoxynucleoside supports. After deprotection, the primary
amine on the C6 analogues is separated from the oligonucleotide by a spacer
arm with a total of 7 -10 atoms and can be labelled or attached to an
enzyme. The C2 analogue is more suitable for the attachment of molecules
designed to react with the oligonucleotide.
Our repertoire of NHS ester derivatives has been expanded to include the NHS-Carboxy-dT-CE Phosphoramidite. By making a dT analog of the Carboxy-Modifier C10, it is possible to label one or multiple sites within an oligonucleotide. This opens up the possibility to label any number of different dyes or molecules within an oligonucleotide when the phosphoramidite is unavailable. Doing so is straightforward and may be done manually off the synthesizer or even in a fully-automated manner on the DNA synthesizer. We have never found conditions which allow the TFA group to be removed from an amino-modifier while the oligonucleotide remains attached to the support. We are able to solve this problem by using a 9-fluorenylmethoxycarbonyl (Fmoc) protecting group. The Fmoc group is removed using a two step procedure, the first to remove the cyanoethyl protection groups and flush the formed acrylonitrile from the synthesis column using 1% diisopropylamine in acetonitrile, and the second to remove the Fmoc group using 10% piperidine in DMF. The amino group so formed on the column can be reacted with a variety of activated esters. We offer Fmoc-Amino-Modifier C6 dT Phosphoramidite as a nucleosidic option and Amino-Modifier Serinol Phosphoramidite as a non-nucleosidic alternative. We also offer S-Bz-Thiol-Modifier C6-dT to join the ranks of thiol-modifiers for oligonucleotide synthesis. Thiol-Modifier C6-dT can be added as usual at the desired locations within a sequence.
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3’-MODIFIERS3’-Amino-Modifier CPGs, containing amino groups protected with the base-labile Fmoc group, are designed to functionalize the 3’-terminus of the target oligonucleotide by the introduction of a primary amine. In an alternative approach, the nitrogen destined to become the 3’-amino group is included in a phthalimide (PT) group which is attached to the support through an amide group attached to the aromatic ring. This simple linkage is very stable to all conditions of oligonucleotide synthesis and contains no chiral center. Using an extended ammonium hydroxide treatment (55°C for 17 hours), the cleavage of the amine from the phthalimide is accomplished along with the deprotection of the oligonucleotide. ABI-style columns are supplied unless otherwise requested.
The 3'-Thiol-Modifier S-S CPG supports are used to introduce 3'-thiol linkages with three and six carbon spacers in oligonucleotides. DTPA CPG is used to introduce a dithiol group at the 3'-terminus. In conjunction with DTPA Phosphoramidite, it is simple to produce oligonucleotides with multiple thiol groups at the 3' terminus, which is ideal for conjugation to gold surfaces. With Glyceryl CPG the 3'-terminus of an oligonucleotide is readily oxidized by sodium periodate to form a 3'-phosphoglycaldehyde. The aldehyde may be further oxidized to the corresponding carboxylic acid. Either the aldehyde or the carboxylate may be used for subsequent conjugation to amine-containing products.
3'-Amino-Modifier C6 dC CPG and 3'-Amino-Modifier C6 dT CPG replace a dC and T, respectively, at the 3'-terminus. Theseproducts allow convenient labelling at the 3' without blocking the terminus from desired enzymatic activity.
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OTHER INSTRUMENT TYPESAll minor bases, RNA products and modifiers are packaged in septum-capped vials suitable for ABI and other instruments. If you would like another type of vial/column add the following to the end of the catalog number.
OTHER INSTRUMENT TYPESAll minor bases, RNA products and modifiers are packaged in septum-capped vials suitable for ABI and other instruments. If you would like another type of vial/column add the following to the end of the catalog number.
INTELLECTUAL PROPERTY3'-Thiol-Modifier 6 S-S CPG was developed by, and is sold under agreement from Berry & Associates. |
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CHEMICAL PHOSPHORYLATIONChemical Phosphorylation Reagent is most commonly used to phosphorylate the 5'-terminus of an oligonucleotide. Although this product is also successful in 3'-phosphorylation, 3'-Phosphate CPG allows direct preparation of oligonucleotides with a 3'-phosphate group. Chemical Phosphorylation Reagent II contains a DMT group on a side chain which is stable to base cleavage and can be left on the oligonucleotide for use in RP purification. The DMT group is later removed with aqueous acid and the side chain is eliminated after brief treatment with aqueous ammonium hydroxide to yield the 5'-phosphate.1 Solid CPR II is similar in performance to CPR II but it is easier to prepare aliquots since it is a powder. Many researchers treat synthesis supports with a hindered base (e.g., diethylamine, diisopropylethylamine, or DBU) post-synthesis to eliminate and remove the cyanoethyl phosphate groups. In this way, the acrylonitrile formed in situ is removed from the support and is not available to alkylate dT residues at the N3 position in the oligos. Since the sulfonylethyl group in 3'-Phosphate CPG is also susceptible to ß-elimination leading to oligo cleavage, this technique is not compatible with 3'-phosphate CPG. Using CPR II CPG, which is base labile but does not support ß-elimination, the cyanoethyl groups can be removed from the oligo prior to cleavage and base deprotection. ABI-style vials and columns are supplied unless otherwise requested.
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INTELLECTUAL PROPERTYChemical Phosphorylation Reagent II is covered by US Patent No.: 5,959,090. (1) A. Guzaev, H.Salo, A. Azhayev, and H. Lonnberg, Tetrahedron, 1995, 51, 9375-9384.
REFERENCE(1) A. Guzaev, H.Salo, A. Azhayev, and H. Lonnberg, Tetrahedron, 1995, 51, 9375-9384. SEE ALSOOTHER INSTRUMENT TYPESAll minor bases, RNA products and modifiers are packaged in septum-capped vials suitable for ABI and other instruments. If you would like another type of vial/column add the following to the end of the catalog number.
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ALDEHYDE MODIFICATIONAldehyde modifiers would be attractive electrophilic substitutions in oligonucleotides since they are able to react with amino groups to form a Schiff's base, with hydrazino groups to form hydrazones, and with semicarbazides to form semi-carbazones. The Schiff's base is unstable and must be reduced with sodium borohydride to form a stable linkage but hydrazones and semicarbazides are very stable linkages. Our collaboration with Epoch Biosciences, a subsidiary of Nanogen, Inc., has allowed us to offer 5'-Aldehyde-Modifier C2 Phosphoramidite. The acetal protecting group is sufficiently hydrophobic for use in RP HPLC and cartridge purification and is readily removed after oligonucleotide synthesis under standard oligonucleotide detritylation conditions with 80% acetic acid / 20% water or 2% aqueous trifluoroacetic acid during cartridge purification. A formylindole nucleoside analogue has been used to introduce aldehyde groups within an oligonucleotide or at the 5' terminus. This product has no protecting group on the aldehyde, which means that deprotection of the modified oligonucleotide can be done without changing preferred conditions.
Tosyl Modification - PRODUCT CURRENTLY UNAVAILABLEUniversal Tosyl Phosphoramidite can be used to insert a tosyl group at the 3' or 5' terminus of an oligonucleotide. The tosyl group may be displaced by nucleophilic substitution using, e.g., a thiol tag, on the synthesis column. Alternatively, the oligonucleotide can be cleaved and deprotected using UltraMild chemistry to yield the fully-deprotected tosyl oligonucleotide. The tosyl group can then be displaced in solution with a protein, antibody, or even an amino-modified nucleic acid of interest.
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INTELLECTUAL PROPERTYThese Products are for research purposes only, and may not be used for commercial, clinical, diagnostic or any other use. The Products are subject to proprietary rights of Epoch Biosciences, Inc. and are made and sold under license from Epoch Biosciences, Inc. There is no implied license for commercial use with respect to the Products and a license must be obtained directly from Epoch Biosciences, Inc. with respect to any proposed commercial use of the Products. “Commercial use” includes but is not limited to the sale, lease, license or other transfer of the Products or any material derived or produced from them, the sale, lease, license or other grant of rights to use the Products or any material derived or produced from them, or the use of the Products to perform services for a fee for third parties (including contract research). A simple agreement must be signed before end-users and custom oligo services may purchase these products for use as defined above. |
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SPACER MODIFIERSThe spacer phosphoramidites C3, 9, C12 and 18 are used to insert a spacer arm in an oligonucleotide. The compounds may be added in multiple additions when a longer spacer is required. 3’-Spacer C3 CPG may also act as a blocker of exonuclease and polymerase activity at the 3’-terminus. dSpacer is used to introduce a stable abasic site within an oligonucleotide. PC Spacer is a photocleavable C3 spacer modifier, part of our line of photocleavable (PC) modifiers
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ALSO SEEPC Modifiers p74 Pyrrolidine p56 OTHER INSTRUMENT TYPESAll minor bases, RNA products and modifiers are packaged in septum-capped vials suitable for ABI and other instruments. If you would like another type of vial/column add the following to the end of the catalog number.
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DendrimersDendrimers are discrete, highly branched, monodispersed polymers that possess patterns reminiscent of the branching of trees. Plain and mixed oligonucleotide dendrimers can be synthesized using novel doubling and trebling phosphoramidite synthons.1,2 Dendrimers offer the following advantages. Incorporation of label using γ-32P-ATP and polynucleotide kinase increases in proportion to the number of 5'-ends. Fluorescent signal also increases in proportion to the number of 5'-ends, if spacers are incorporated between the labels and the ends of the branches. When using a dendrimeric oligonucleotide as a PCR primer, the strand bearing the dendrimer is resistant to degradation by T7 Gene 6 exonuclease making it easy to convert the double-stranded product of the PCR to a multiply labelled, single-stranded probe. Enhanced stability of DNA dendrimers makes them useful as building blocks for the ‘bottom up' approach to nano-assembly. These features also suggest applications in DNA chip technology when higher temperatures are required, for example, to melt secondary structure in the target.
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REFERENCES(1) M.S. Shchepinov, I.A. Udalova, A.J. Bridgman, and E.M. Southern, Nucleic Acids Res, 1997, 25, 4447-4454. (2) M.S. Shchepinov, K.U. Mir, J.K. Elder, M.D. Frank-Kamenetskii, and E.M. Southern, Nucleic Acids Res, 1999, 27, 3035-41. (3) T. Horn, C.A. Chang, and M.S. Urdea, Nucleic Acids Res, 1997, 25, 4842-4849. INTELLECTUAL PROPERTYDoublers and Trebler are supplied under license from ISIS Innovation Limited. OTHER INSTRUMENT TYPESAll minor bases, RNA products and modifiers are packaged in septum-capped vials suitable for ABI and other instruments. If you would like another type of vial/column add the following to the end of the catalog number. |
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BRANCHING PHOSPHORAMIDITEA branching monomer is required to construct comb-like oligonucleotide probes. The developers of the comb system from Chiron Corporation evaluated3 several protecting groups for the branch point and chose levulinyl (LEV), which is specifically removed using a reagent containing hydrazine hydrate, acetic acid and pyridine.
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Photocleavable MonomersPC Biotin Phosphoramidite can be used to prepare 5’-biotinylated oligonucleotides suitable for capture by streptavidin in a mode similar to our popular 5’ Biotin Phosphoramidite. Amino- and thiol-modified oligonucleotides have proven to be very useful for the attachment of a variety of haptens and fluorophores, as well as for the tethering of the oligonucleotides to a diversity of beads and surfaces. PC Amino-Modifier Phosphoramidite is used to prepare 5’-amino-modified oligonucleotides suitable for subsequent photocleavage. PC Spacer Phosphoramidite can be used as an intermediary to attach any modification reagent, available as a phosphoramidite, to the terminus of oligonucleotides. After photocleavage, a 5’-phosphate is generated on the DNA, rendering it suitable for further biological transformations, such as gene construction and cloning after ligation. A versatile photocleavable DNA building block has been described by researchers in Washington University, Missouri and used in phototriggered hybridization.1 This reagent has also been used in the design of multifunctional DNA and RNA conjugates2 for the in vitro selection of new molecules catalyzing biomolecular reactions. Researchers at Bruker Daltonik in Germany have also developed genoSNIP, a method for single-nucleotide polymorphism (SNP) genotyping by MALDI-TOF mass spectrometry.3 This method uses size reduction of primer extension products by incorporation of the photocleavable linker for phototriggering strand breaks near to the 3' end of the extension primer. PC Linker can be incorporated into oligonucleotides at any position by standard automated DNA synthesis methodology. PC Linker Phosphoramidite has the added advantage in that photocleavage results in monophosphate fragments at both the 3'- and 5'-termini of the oligonucleotide fragments.
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INTELLECTUAL PROPERTYGlen Research offers PC Biotin, PC Amino-Modifier and PC Spacer products in association with AmberGen, Inc. and Link Technologies, Ltd. For a commercial application license, please contact AmberGen, Inc., 617-923-9990 (sales@ambergen.com), http://www.ambergen.com. SEE ALSOReferences(1) P. Ordoukhanian and J-S. Taylor, J. Am. Chem. Soc., 117, 9570-9571, 1995. (2a) F. Hausch and A. Jäschke, Nucleic Acids Research, 2000, 28, e35. (3) T. Wenzel, T. Elssner, K. Fahr, J. Bimmler, S. Richter, I. Thomas, and M. Kostrzewa, Nucleosides, Nucleotides & Nucleic Acids, 2003, 22, 1579-1581. |
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OLIGONUCLEOTIDE-PEPTIDE CONJUGATESThe OPeC™ method for the synthesis of Oligonucleotide-Peptide Conjugates, originally described by Stetsenko and Gait1,2, is based on the “native ligation” of an N-terminal thioester-functionalized peptide to a 5'-cysteinyl oligonucleotide. OPeC™reagents have been developed by Link Technologies Ltd. In OPeC™ conjugation strategies, peptide and oligonucleotide units are easily assembled separately on their own supports using conventional synthesizers and methodology. Each biomolecule is designed to carry a reactive functionality that is released upon full deprotection and cleavage from the support. The components can then be conjugated in aqueous/organic solution by selective reaction of these functionalities.
| References(1) D.A. Stetsenko and M.J. Gait, J Org Chem, 2000, 65, 4900-4908. (2) D.A. Stetsenko and M.J. Gait, Nucleos Nucleot Nucleic Acids, 2000, 19, 1751-1764. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Conjugation using Click ChemistryThe copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) reaction between azides and alkynes to form 1,2,3-triazoles, as reported1 by Sharpless, was found to be so exquisitely regioselective and efficient at even the most mild conditions that Sharpless coined the term ‘Click Chemistry' to describe it. The use of this method for DNA modification has been somewhat delayed by the fact that copper ions damage DNA, typically yielding strand breaks.2 As these problems have now been overcome by the use of copper(I)-stabilizing ligands (e.g., tris(benzyltriazolylmethyl)amine, TBTA3), Carell et al. and Seela et al. discovered that the CuAAC reaction can be used to functionalize alkyne-modified DNA nucleobases with extremely high efficiency.4 Oligonucleotides bearing a single nucleosidic alkyne group can be prepared using a C8-Alkyne-dC or dT-CE Phosphoramidite. Purified oligonucleotides are usually modified with 2-5 equivalents of the corresponding marker-azide (e.g., fluorescent-dye azides). After the addition of precomplexed Cu(I), complete conversion to the labelled oligo is observed in a time span between 30 min and 4 hours. After a simple precipitation step, labelled oligonucleotides can be recovered in near quantitative yields. Using a combination of C8-Alkyne, C8-TIPS-Alkyne and C8-TMS-Alkyne, it is possible to label oligonucleotides in up to three separate click reactions. The alkyne groups on the last two monomers are protected, respectively, with triisopropylsilyl (TIPS) and trimethylsilyl (TMS) protecting groups.5,6 The first click reaction on solid phase on a C8-Alkyne yields the singly modified oligonucleotide with full retention of the TIPS and/or TMS protecting group. For double click, a C8-TIPS-Alkyne is used as the second nucleoside and the TIPS protecting group is cleaved with tetrabutylammonium fluoride (TBAF) without causing any damage to the DNA. The second click reaction in solution yields the doubly modified oligonucleotide in excellent yield. For the introduction of three different labels, all three nucleosides are introduced into oligonucleotides. The first click reaction is performed directly on the resin. The singly modified oligonucleotide is subsequently cleaved from the support with concomitant cleavage of the TMS group and retention of the TIPS protecting group. The second click reaction is performed in solution. Precipitation of the doubly modified oligonucleotide, cleavage of the TIPS group with TBAF, and a subsequent third click reaction in solution furnishes the desired triply modified oligonucleotide in excellent overall yield.
OLIGO-CLICK KITSOligo-Click Kits contain an air-stable, insoluble Cu(I) source in pellet form in a pre-loaded and ready-to-use vial. Within the kit, the TBTA ligand is replaced by an activator which is compatible with both aqueous and organic solvents. This innovative combination of catalyst and ligand/activator results in a much easier labeling work-flow of only three simple steps. The preparation of the oligonucleotide labeling via CuAAC now requires only a minimal hands-on time of a few minutes or even less and can be carried out in air without any additional precautions. Glen Research is offering the following kits in collaboration with baseclick GmbH.
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References[1] C.W. Tornoe, C. Christensen, M. Meldal, J. Org. Chem. 2002, 67, 3057-3064; V. V. Rostovtsev, L. G. Green, V. V. Fokin, K. B. Sharpless, Angew. Chem. 2002, 114, 2708-2711; Angew. Chem. Int. Ed. 2002, 41, 2596-2599. INTELLECTUAL PROPERTY All products of baseclick are patent protected and available in collaboration with baseclick. Baseclick GmbH holds a worldwide license for the research market of the “Click Chemistry” patent from “The Scripps Research Institute”: | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Conjugation using Click Chemistry (CONT.)Oligonucleotides prepared using 5'-Hexynyl Phosphoramidite are stable to standard deprotection conditions and exhibit a slightly increased retention time on RP HPLC. Azides are not compatible with oligonucleotide synthesis using phosphoramidites so a post-synthesis reaction is required. Azidobutyrate NHS Ester is used1 for azido-modification of amines at either the 3'-end or the 5'-end of an oligo and it can even be used for internal modification on an Amino-Modifier-C6 dX residue within the sequence. Specific to the 5'-terminus, 5'-Bromohexyl Phosphoramidite is added in the last cycle. This modifier can then be easily transformed into a 5'-azido group by displacement of bromide using sodium azide.2 Alkyne NHS ester allows the functionalization of an amino moiety in a variety of molecules, including DNA and RNA oligonucleotides as well as peptides or proteins. We also offer two products for use in Click Chemistry based upon our 1,3-diol product portfolio with the serinol backbone - a phosphoramidite for adding an alkyne group at the 5' terminus or within the sequence, and a synthesis support for labelling the 3' terminus of oligonucleotides with an alkyne group. A 5'-iodo-modified oligonucleotide (prepared using 5'-Iodo-dT) can be quantitatively converted to the corresponding 5'-azide.
Copper-free Click ChemistryAt Glen Research, our goal was to offer a copper-free click phosphoramidite reagent with the following properties:
From the variety of cyclooctyne-based copper-free click reagents so far described, we have chosen to offer compounds based on a dibenzo-cyclooctyne (DBCO) structure. We are offering 5’-DBCO-TEG Phosphoramidite for preparing oligos with a 5’-DBCO modification and DBCO-dT-CE Phosphoramidite for inserting a DBCO group at any position within the oligonucleotide. DBCO-sulfo-NHS Ester is also offered for post-synthesis conjugation reactions. DBCO-modified oligos may be conjugated with azides in organic solvents, such as DMSO, or aqeous buffers. Depending on the azide used, the reaction will go to completion in 4-17 hours at room temperature. Simple desalting on a Glen Gel-Pak™ leads to a product with virtually quantitative conjugation efficiency.
Conjugation using Click Chemistry (CONT.)Glen Research is collaborating with baseclick Gmbh to offer a variety of interesting alkyne phosphoramidites. We offer some azide products and are now embarking on an expansion of our azide catalog. Our strategy is to offer first our most popular labels for general interest and, subsequently, we will add azide products that are not compatible with phosphoramidite chemistry. Biotin is still our most commonly used label and biotinTEG, with its hydrophilic triethylene glycol spacer, is the most popular biotin product. Desthiobiotin is a biotin analogue that is well captured by streptavidin but the captured product can be easily released by applying a biotin solution to the streptavidin beads. 6-FAM is our most popular fluorescein derivative and we offer azides of both 6-FAM and pivaloyl-protected 6-FAM for situations where subsequent reactions require the 6-FAM to be protected. In both 6-FAM products, the hydrophilic TEG spacer is again used. The azides are offered in 25 and 100 µmole packs for convenient oligonucleotide labelling. 7-Hydroxycoumarin, also known as umbelliferone, is a highly fluorescent, pH-sensitive fluorophore that emits in the blue region of the spectrum. However, its fluorescence is strongly quenched if the hydroxyl is alkylated or phosphorylated, making it useful in high-throughput screening for phosphatases and lipases. Interestingly, it was found that the 3-azido derivative is also highly quenched but, upon reaction with an alkyne in the presence of copper to form the triazole, the fluorescence is restored.1 The clicked coumarin emits at a lambda max of 480 nm and absorbs at 358 nm. HEX and TET are two of our most popular fluorescein-based dyes for labelling oligonucleotides. We are happy to offer 6-HEX and 6-TET Azides for use in click conjugations.
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References(1) R. Kumar, et al., Journal of the American Chemical Society, 2007, 129, 6859-6864. STABILITY NOTESOligonucleotides containing a 5'-iodo group are prepared conventionally with the exception that deprotection is carried out in ammonium hydroxide at room temperature for 24 hours. Under these conditions, degradation of the iodo group was less than 2%. See Also |
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