TERMINUS MODIFIERS

Glen 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.

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.

SEQUENCE MODIFIERS

Sequence 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.

Item	Catalog No.	Pack	Price ($)
Amino-Modifier C6 dA	10-1089-90	100 µmole	205.00
	10-1089-02	0.25g	455.00
Amino-Modifier C6 dC	10-1019-90	100 µmole	225.00
	10-1019-02	0.25g	450.00
Amino-Modifier C6 dG	10-1099-95	50 µmole	240.00
	10-1099-90	100 µmole	480.00
	10-1099-02	0.25g	1100.00
Carboxy-dT	10-1035-90	100 µmole	180.00
	10-1035-02	0.25g	360.00
Amino-Modifier C2 dT	10-1037-90	100 µmole	180.00
	10-1037-02	0.25g	360.00
	10-1037-05	0.5g	720.00
Amino-Modifier C6 dT	10-1039-90	100 µmole	180.00
	10-1039-02	0.25g	360.00
	10-1039-05	0.5g	720.00
Fmoc-Amino-Modifier C6 dT-CE Phosphoramidite	10-1536-90	100 µmole	180.00
	10-1536-02	0.25g	360.00

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.

3’-MODIFIERS

3’-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.

Item	Catalog No.	Pack	Price ($)
3'-Amino-Modifier C7 CPG 500	20-2957-01	0.1g	95.00
	20-2957-10	1.0g	675.00
1 µmole columns	20-2957-41	Pack of 4	140.00
0.2 µmole columns	20-2957-42	Pack of 4	85.00
10 µmole column (ABI)	20-2957-13	Pack of 1	250.00
15 µmole column (Expedite)	20-2957-14	Pack of 1	375.00
3'-Amino-Modifier C7 CPG	20-2958-01	0.1g	95.00
	20-2958-10	1.0g	675.00
1 µmole columns	20-2958-41	Pack of 4	140.00
0.2 µmole columns	20-2958-42	Pack of 4	85.00
10 µmole column (ABI)	20-2958-13	Pack of 1	250.00
15 µmole column (Expedite)	20-2958-14	Pack of 1	375.00
3’-Amino-Modifier Serinol CPG	20-2997-01	0.1g	95.00
	20-2997-10	1.0g	675.00
0.2 µmole columns	20-2997-42	Pack of 4	85.00
1 µmole columns	20-2997-41	Pack of 4	140.00
10 µmole column (ABI)	20-2997-13	Pack of 1	250.00
15 µmole column (Expedite)	20-2997-14	Pack of 1	375.00
3'-PT-Amino-Modifier C3 CPG	20-2954-01	0.1g	95.00
	20-2954-10	1.0g	675.00
1 µmole columns	20-2954-41	Pack of 4	140.00
0.2 µmole columns	20-2954-42	Pack of 4	85.00
10 µmole column (ABI)	20-2954-13	Pack of 1	250.00
15 µmole column (Expedite)	20-2954-14	Pack of 1	375.00
3'-PT-Amino-Modifier C6 CPG	20-2956-01	0.1g	95.00
	20-2956-10	1.0g	675.00
1 µmole columns	20-2956-41	Pack of 4	140.00
0.2 µmole columns	20-2956-42	Pack of 4	85.00
10 µmole column (ABI)	20-2956-13	Pack of 1	250.00
15 µmole column (Expedite)	20-2956-14	Pack of 1	375.00
3'-PT-Amino-Modifier C6 PS
200 nmole columns (ABI 3900)	26-2956-52	Pack of 10	220.00
40 nmole columns (ABI 3900)	26-2956-55	Pack of 10	220.00

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.

Item	Catalog No.	Pack	Price ($)
3'-Thiol-Modifier C3 S-S CPG	20-2933-01	0.1g	85.00
	20-2933-10	1.0g	600.00
1 µmole columns	20-2933-41	Pack of 4	125.00
0.2 µmole columns	20-2933-42	Pack of 4	75.00
10 µmole column (ABI)	20-2933-13	Pack of 1	225.00
15 µmole column (Expedite)	20-2933-14	Pack of 1	350.00
3’-Thiol-Modifier C6 S-S CPG	20-2936-01	0.1g	85.00
	20-2936-10	1.0g	600.00
1 µmole columns	20-2936-41	Pack of 4	125.00
0.2 µmole columns	20-2936-42	Pack of 4	75.00
10 µmole column (ABI)	20-2936-13	Pack of 1	225.00
15 µmole column (Expedite)	20-2936-14	Pack of 1	350.00
3’-DTPA CPG	20-2937-01	0.1g	125.00
	20-2937-10	1.0g	895.00
1 µmole columns	20-2937-41	Pack of 4	185.00
0.2 µmole columns	20-2937-42	Pack of 4	110.00
10 µmole column (ABI)	20-2937-13	Pack of 1	335.00
15 µmole column (Expedite)	20-2937-14	Pack of 1	520.00
3'-Glyceryl CPG	20-2902-01	0.1g	85.00
	20-2902-10	1.0g	600.00
1 µmole columns	20-2902-41	Pack of 4	125.00
0.2 µmole columns	20-2902-42	Pack of 4	75.00
10 µmole column (ABI)	20-2902-13	Pack of 1	225.00
15 µmole column (Expedite)	20-2902-14	Pack of 1	350.00
3'-Amino-Modifier C6 dC CPG	20-2019-01	0.1g	120.00
	20-2019-10	1.0g	995.00
1 µmole columns	20-2019-41	Pack of 4	200.00
0.2 µmole columns	20-2019-42	Pack of 4	120.00
10 µmole column (ABI)	20-2019-13	Pack of 1	300.00
15 µmole column (Expedite)	20-2019-14	Pack of 1	450.00
3'-Amino-Modifier C6 dT CPG	20-2039-01	0.1g	96.00
	20-2039-10	1.0g	800.00
1 µmole columns	20-2039-41	Pack of 4	160.00
0.2 µmole columns	20-2039-42	Pack of 4	96.00
10 µmole column (ABI)	20-2039-13	Pack of 1	240.00
15 µmole column (Expedite)	20-2039-14	Pack of 1	360.00

CHEMICAL PHOSPHORYLATION

Chemical 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 ABI-style vials and columns are supplied unless otherwise requested.

Item	Catalog No.	Pack	Price ($)
Chemical Phosphorylation Reagent	10-1900-90	100 µmole	50.00
	10-1900-02	0.25g	160.00
3'-Phosphate CPG	20-2900-01	0.1g	70.00
	20-2900-10	1.0g	480.00
1 µmole columns	20-2900-41	Pack of 4	100.00
0.2 µmole columns	20-2900-42	Pack of 4	60.00
10 µmole column (ABI)	20-2900-13	Pack of 1	180.00
15 µmole column (Expedite)	20-2900-14	Pack of 1	280.00
3'-Phosphate PS
200 nmole columns (ABI 3900)	26-2900-52	Pack of 10	150.00
40 nmole columns (ABI 3900)	26-2900-55	Pack of 10	150.00
3'-Phosphate CPG (High Load)	25-2900-01	0.1g	85.00
	25-2900-10	1.0g	600.00
2.5 µmole columns	25-2900-46	Pack of 4	120.00
Chemical Phosphorylation Reagent II	10-1901-90	100 µmole	60.00
(CPR II)	10-1901-02	0.25g	200.00
Solid Chemical Phosphorylation Reagent II	10-1902-90	100 µmole	60.00
(Solid CPR II)	10-1902-02	0.25g	200.00

ALDEHYDE MODIFICATION

Aldehyde 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.

Item	Catalog No.	Pack	Price ($)
5'-Aldehyde-Modifier C2 Phosphoramidite	10-1933-90	100 µmole	85.00
	10-1933-02	0.25g	325.00
Formylindole CE Phosphoramidite	10-1934-90	100 µmole	85.00
	10-1934-02	0.25g	325.00

SPACER MODIFIERS

The 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

Item	Catalog No.	Pack	Price ($)
Spacer Phosphoramidite 9	10-1909-90	100 µmole	75.00
	10-1909-02	0.25g	240.00
Spacer Phosphoramidite C3	10-1913-90	100 µmole	75.00
	10-1913-02	0.25g	240.00
dSpacer CE Phosphoramidite	10-1914-90	100 µmole	85.00
	10-1914-02	0.25g	295.00
Pyrrolidine-CE Phosphoramidite	10-1915-95	50 µmole	190.00
(PYR)	10-1915-90	100 µmole	380.00
	10-1915-02	0.25g	1085.00
Spacer Phosphoramidite 18	10-1918-90	100 µmole	95.00
	10-1918-02	0.25g	240.00
Spacer C12 CE Phosphoramidite	10-1928-90	100 µmole	95.00
	10-1928-02	0.25g	240.00
3'-Spacer C3 CPG	20-2913-01	0.1g	70.00
	20-2913-10	1.0g	480.00
1 µmole columns	20-2913-41	Pack of 4	100.00
0.2 µmole columns	20-2913-42	Pack of 4	60.00
10 µmole column (ABI)	20-2913-13	Pack of 1	180.00
15 µmole column (Expedite)	20-2913-14	Pack of 1	280.00
PC Spacer Phosphoramidite	10-4913-90	100 µmole	135.00
	10-4913-02	0.25g	395.00

Dendrimers

Dendrimers 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.

Item	Catalog No.	Pack	Price ($)
Symmetric Doubler Phosphoramidite	10-1920-90	100 µmole	150.00
	10-1920-02	0.25g	240.00
Asymmetric Doubler Phosphoramidite	10-1921-90	100 µmole	180.00
	10-1921-02	0.25g	300.00
Trebler Phosphoramidite	10-1922-90	100 µmole	180.00
	10-1922-02	0.25g	300.00
Long Trebler Phosphoramidite	10-1925-90	100 µmole	200.00
	10-1925-02	0.25g	300.00

BRANCHING PHOSPHORAMIDITE

A 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.  

Item	Catalog No.	Pack	Price ($)
5-Me-dC Brancher-CE Phosphoramidite	10-1018-90	100 µmole	205.00
	10-1018-02	0.25g	505.00

Photocleavable Monomers

PC 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.

OLIGONUCLEOTIDE-PEPTIDE CONJUGATES

The 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.

Item	Catalog No.	Pack	Price ($)
Oligonucleotide-Peptide Conjugation Kit (OPeC™)	50-1000-01	each	495.00
Each OPeC kit consists of the following components, which can also be ordered individually as shown:
Oligo Modifying Reagent	50-1001-01	150µmole	290.00
Peptide Modifying Reagent	50-1002-01	450µmole	250.00
TCEP (Conjugation Reagent)	50-1003-01	230mg	75.00

Conjugation using Click Chemistry

The conjugation reaction between azides and alkynes to form 1,2,3-triazoles, as reported³ 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. Oligonucleotides prepared using 5’-Hexynyl Phosphoramidite are stable to standard deprotection conditions and exhibit a slightly increased retention time on RP HPLC. A number of azide labels are commercially available for click conjugation to a 5’-hexynyl-modified oligonucleotide. Azides are not compatible with oligonucleotide synthesis using phosphoramidites so a post-synthesis reaction is likely to be required. We found we were able to quantitatively convert a 5’-iodo-modified oligonucleotide (prepared using 5’-Iodo-dT) to the corresponding 5’-azide.