Duplex Stabilization (Catalog as PDF)

c-5 propyne derivatives and G-Clamp

Substitution of C-5 propynyl-dC (pdC) for dC and C-5 propynyl-dU (pdU) for dT are effective strategies to enhance base pairing. Using these base substitutions, duplex stability and melting temperatures are raised by the following amounts: C-5 propynyl-C 2.8° per substitution; C-5 propynyl-U 1.7° per substitution. AP-dC (G-clamp) substitutes for dC and is another very important modified nucleoside that enhances hybridization by 7-21° per substitution depending upon the sequence and location of the AP-dC. The ability of these modified bases to enhance binding while maintaining specificity has proven useful in antisense research and in the synthesis of high affinity probes. AP-dC is also a fluorescent nucleoside and should find uses in DNA structural research.
Item Catalog No. Pack Price ($)
10-1014-90 100µm 85.00
10-1014-02 0.25g 245.00
10-1014-05 0.5g 490.00
10-1054-90 100µm 65.00
10-1054-02 0.25g 195.00
10-1054-05 0.5g 390.00
10-1097-95 50µm 230.00
  (G-Clamp) 10-1097-90 100µm 460.00
10-1097-02 0.25g 1175.00
pdC
pdC
pdU
pdU
AP-dC
AP-dC

Intellectual Property

AP-dC is covered by patents or patents pending owned by Ionis Pharmaceuticals, Inc. ("Ionis"). Purchase of this product includes a limited license to use this product solely as an intermediary in the manufacture of oligomers containing 6 or more nucleosides ("Oligonucleotides") for internal research. This license specifically excludes (and you have no right to use this product or Oligonucleotides for): (a) therapeutic or diagnostic applications (including products or services that incorporate this product or Oligonucleotides), (b) any in vivo toxicity/safety study in support of an investigational new drug application (or foreign counterpart), (c) resale (including sale of any products or services that incorporate this product or Oligonucleotides) or (d) gene functionalization activities (including products or services that incorporate data derived from gene functionalization activities) if such activities have commercial application, any and all of which require a separate license from Ionis. Neither this product nor any product created through its use may be used in human clinical trials.

A simple agreement must be signed before end-users and custom
oligo services may purchase this product for use as defined above. http://www.glenresearch.com/ Reference/AP-dCEmail.pdf

bases affecting duplex stAbility

C-5 methyl pyrimidine nucleosides are known to stabilize duplexes relative to the non-methylated bases.  Therefore, enhanced binding can be achieved using 5-methyl-dC in place of dC, duplex melting temperature being increased by 1.3°.  Ac-5-Me-dC-CE Phosphoramidite is fully compatible with AMA deprotection and none of the N4-Me transamination mutant is observed on deprotection.  2,6-Diaminopurine 2'-deoxyriboside (2-amino-dA) forms an additional hydrogen bond with Thymidine, thereby leading to duplex stabilization with a melting temperature increase of 3°.  Our 2-amino-dA monomer exhibits fast and effective deprotection in ammonium hydroxide and it is stabilized to depurination during synthesis.  Note: we recommend the use of 0.5 M CSO in anhydrous acetonitrile (40-4632-xx) for best results with multiple additions of 2-amino-dA.
Item Catalog No. Pack Price ($)
10-1060-90 100µm 50.00
10-1060-02 0.25g 120.00
10-1560-90 100µm 50.00
10-1560-02 0.25g 120.00
5-Me-dC
5-Me-dC
Ac-5-Me-dC
Ac-5-Me-dC

OTHER INSTRUMENT TYPES

All minor bases, RNA products and modifiers are packaged in septumcapped 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.

Monomers  

For Instrument type
Add
Expedite
E
Mermade
M

Columns  

For Instrument type
Add
Expedite
E
Applied Biosystems 3900
A
Mermade
M

Please inquire for availability of columns for other instrument types.

bases affecting duplex stAbility (Part 2)

The simplest approach to the design of high affinity primers and probes is to substitute A sites with 2-amino-A, since the 2-amino-A-T base pair is equivalent in strength to the G-T base pair. 2-Amino-A also destabilizes A-G wobble mismatches, thus increasing specificity. In 1998, we introduced a 2-amino-dA monomer which exhibits fast and effective deprotection in ammonium hydroxide and it is stabilized to depurination during synthesis. We now recommend the use of 0.5 M CSO in anhydrous acetonitrile (40-4632-xx) for best results with multiple additions of 2-amino-dA. This is because the bis formamidine protected 2-amino-dA leads to significant strand scission when standard iodine oxidation is used during synthesis. For this reason, we have also added Pac-2-Amino-dA, a monomer with optimized protection to meet the following criteria: stable during oligonucleotide synthesis, oxidation, and detritylation; labile towards common deprotection conditions (NH3, AMA, MeNH2); and the nucleobase protecting groups are cleaved under fairly mild conditions.
Item Catalog No. Pack Price ($)
10-1085-95 50µm 70.00
  (2,6-diaminopurine) 10-1085-90 100µm 125.00
10-1085-02 0.25g 250.00
(2,6-diaminopurine)
10-1585-95 50µm 70.00
10-1585-90 100µm 125.00
10-1585-02 0.25g 250.00
2-Amino-dA
2-Amino-dA
Pac-2-Amino-dA
Pac-2-Amino-dA

OTHER INSTRUMENT TYPES

All minor bases, RNA products and modifiers are packaged in septumcapped 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.

Monomers  

For Instrument type
Add
Expedite
E
Mermade
M

Columns  

For Instrument type
Add
Expedite
E
Applied Biosystems 3900
A
Mermade
M

Please inquire for availability of columns for other instrument types.

See Also

bases affecting duplex stAbility (Part 2)

Sequences with high GC content may contain mismatches and still hybridize because of the high stability of the G-C base pair. The N4-ethyl analogue of dC (N4-Et-dC) hybridizes specifically to natural dG but the stability of the base pair is reduced to about the level of an AT base pair.

Coupling N6-Me-dA (10-1003) and N4-Et-dC (10-1068) with 1H-tetrazole leads to a trace of branching at the secondary amine positions, while DCI leads to around 15% branching. In collaboration with Berry and Associates, the acetyl protected monomers were prepared. Acetyl protection was chosen since it would block branching reactions. Oligonucleotides synthesized using these monomers proved to be compatible with all popular deprotection strategies from UltraMild to UltraFast. When the acetyl protected monomers were compared with the unprotected monomers using DCI as activator, branching was reduced from 15% to zero.
Item Catalog No. Pack Price ($)
10-1068-95 50µm 125.00
10-1068-90 100µm 225.00
10-1068-02 0.25g 675.00
10-1513-95 50µm 125.00
10-1513-90 100µm 225.00
10-1513-02 0.25g 675.00
10-1003-90 100µm 162.50
10-1003-02 0.25g 495.00
10-1503-90 100µm 162.50
10-1503-02 0.25g 495.00
N4-Et-dC
N4-Et-dC
N4-Ac-N4-Et-dC
N4-Ac-N4-Et-dC
N6-Me-dA
N6-Me-dA
N6-Ac-N6-Me-dA
N6-Ac-N6-Me-dA

OTHER INSTRUMENT TYPES

All minor bases, RNA products and modifiers are packaged in septumcapped 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.

Monomers  

For Instrument type
Add
Expedite
E
Mermade
M

Columns  

For Instrument type
Add
Expedite
E
Applied Biosystems 3900
A
Mermade
M

Please inquire for availability of columns for other instrument types.

Zip Nucleic Acids (ZNA®)

Spermine phosphoramidite is used to produce oligospermine-oligonucleotide conjugates - Zip Nucleic Acids (ZNA®) Oligos. The name reflects the presumed mode of action. The conjugates are believed to use the oligospermine to seek out and move along (scan) oligonucleotide strands until the probe complementary sequence is located. The oligospermine then performs the function of stabilizing the formed duplex by reducing electrostatic repulsion, thereby leading to significantly increased binding affinities. ZNA® Oligos have found use in the following applications: Multiplex PCR; PCR of AT-rich Regions; RT qPCR; Detection of MicroRNA; Improved SNP Discrimination; and Antisense and Antigene Effects. Spermine phosphoramidite is simple to use in oligonucleotide synthesis and can be added multiple times at the 3' or 5' terminus. Deprotection and isolation are also straightforward. HPLC analysis of the conjugates requires high pH to suppress the ionization of the spermine residues.
Item Catalog No. Pack Price ($)
10-1939-95 50µm 145.00
10-1939-90 100µm 270.00
10-1939-02 0.25g 525.00
Spermine
Spermine

Intellectual Property

"Spermine phosphoramidite" synthon is the subject matter of U.S. Divisional Patent Application 14/745,871, European Patent No. EP 1973927 B1 and foreign equivalents for which Polyplus-transfection is the co-owner. Product is sold for research purposes only. Product shall not be used to manufacture oligospermine oligonucleotide conjugates for use in diagnostics, clinical or commercial applications including use in humans. There is no implied license to manufacture oligospermine oligonucleotide conjugates for diagnostic, clinical or commercial applications, including but not limited to contract research. Please contact Polyplus-transfection at licensing@polyplus-transfection.com to obtain a license for such use.

ZNA® is a registered trademark of Polyplus-transfection SA.

Caps for Increased Duplex Stability and Base-Pairing Fidelity at Termini

New cap structures allow for the preparation of hybridization probes with increased affinity for complementary sequences. The monomers used to prepare capped oligonucleotides are phosphoramidites that can be readily introduced via automated DNA synthesis at the end of solid phase syntheses. The caps favor the formation of stable Watson-Crick duplexes by stacking on the terminal base pair (Figures 1 and 2).

Fig1 fig2
FIGURE 1: STACKING OF CAP ON 5' TERMINAL BASE PAIRFIGURE 2:
STACKING OF Uaq CAP ON 3' TERMINAL BASE PAIR


Melting point increases of over 10°C per modification can be realized for short duplexes.1,2 The caps fit canonical Watson-Crick base pairs and do not stack well on mismatched base pairs. This leads to increased base pairing selectivity at the terminal and the penultimate position of oligonucleotides featuring the caps. Base pairing fidelity is usually low at the termini, where fraying occurs frequently in the absence of caps. The beneficial effects of the caps are also realized when longer target strands are bound, so there is no need for blunt ends for the duplexes formed.1,2 The caps, when attached to the 5' terminus of an oligonucleotide, also facilitate purification as their lipophilicity leads to prolonged retention on reversed phase columns or cartridges. Finally, capping of termini may discourage the degradation of oligonucleotides by exonucleases.

3'-Uaq Cap CPG, a Uridine support modified with a 2'-anthraquinone residue, is the most effective oligonucleotide cap known to date.3,4 For short hybrid duplexes between DNA probes and RNA target strands, the increase in Tm is up to 18°C and the modification is effective in increasing the Tm of DNA:DNA, RNA:RNA, and DNA:RNA hybrid duplexes. 3'-Uaq Cap also increases probe specificity by depressing the melting point of terminal mismatches.
Item Catalog No. Pack Price ($)
10-1986-90 100µm 195.00
10-1986-02 0.25g 495.00
10-1987-90 100µm 195.00
10-1987-02 0.25g 495.00
20-2980-01 0.1g 180.00
20-2980-10 1.0g 1500.00
  1 µmole columns 20-2980-41 Pack of 4 300.00
  0.2 µmole columns 20-2980-42 Pack of 4 150.00
  10 µmole column (ABI) 20-2980-13 Pack of 1 750.00
  15 µmole column (Expedite) 20-2980-14 Pack of 1 1125.00
5'-Trimethoxystilbene Cap
5'-Trimethoxystilbene Cap
5'-Pyrene Cap
5'-Pyrene Cap
3'-Uaq Cap CPG
3'-Uaq Cap CPG

References

  1. Dogan, Z.; Paulini, R.; Rojas Stütz, J. A.; Narayanan, S.; Richert, C. J. Amer. Chem. Soc. 2004, 126, 4762- 4763.
  2. Narayanan, S.; Gall, J.; Richert, C. Nucleic Acids Res. 2004, 32, 2901- 2911.
  3. A. Patra, C. Richert, J. Amer. Chem. Soc., 2009, 131, 12671-12681.
  4. C. Ahlborn, K. Siegmund, C. Richert, J. Amer. Chem. Soc., 2007, 129, 15218-15232.

Contact to place an order.
INTERNATIONAL USERS: Contact your distributor or order from USA
Contact to access more than 50 years of oligo expertise for help in designing oligos for your unique application.
Glen Research Corporation, Copyright © 2019