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.
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:
• 5'-Amino-Modifier C6-PDA
• Hydrophobic 5'-Amino-Modifier C12-PDA
• Hydrophilic 5'-Amino-Modifier-TEG-PDA
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.
Amino-Modifier dA, Amino-Modifier dC, N2-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.
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.
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
3'-Amino-Modifier C6 dC CPG and 3'-Amino-Modifier C6 dT CPG replace a dC and T, respectively, at the 3'-terminus. These products allow convenient labelling at the 3' without blocking the terminus from desired enzymatic activity.