2′-O,4′-C-Methylene Bridged Nucleic Acid Modification Promotes Pyrimidine Motif Triplex DNA Formation at Physiological pH

The potential for gene repair via triple helix formation

“…RNA third strands formed more stable pyrimidinemotif triplexes than the corresponding DNA strands (42), which prompted the use in third strands of 2′-OMethoxy (OMe) sugar residues (27,43), and recently, 2′, 4′ bridged ribose substitutions (44). These modifications preorganize the third strand in a conformation that is compatible with triplex formation and imposes minimal distortion on the underlying duplex (28,44).…”

Chemical Tools for Targeted Mutagenesis of DNA Based on Triple Helix Formation

“…A number of chemical modifications of TFOs such as to the nucleobase moiety or to the sugar backbone has been attempted to improve the binding affinity of the TFOs. 59,60) However, there is no wellestablished assay system for the evaluation of triplex stability in vivo. Inhibition of transcription with TFOs may include factors other than triple helix formation and may not be simple reflections of triplex stability in vivo.…”

Chemical Modification of Oligonucleotides: A Novel Approach Towards Gene Targeting

“…Kinetic experiments for the triplex formation were performed on a BIACORE J instrument (GE Healthcare, U.S.A.), in which a real-time biomolecular interaction was measured with a laser biosensor, essentially as described previously [12,13,[19][20][21][22][23][24][25]27]. The layer of a SA sensor tip with immobilized streptavidin was equilibrated with 10 mM sodium cacodylatecacodylic acid at pH 6.8 containing 200 mM NaCl and 20 mM MgCl2 at a flow rate of 30 µl/min.…”

“…Isothermal titration experiments for the triplex formation were carried out on a VP ITC system (Microcal Inc., U.S.A.), essentially as described previously [12,13,[19][20][21][22][25][26][27]. The TFO (Figure 1c) and Pur23A•Pyr23T duplex (Figure 1c) solutions were prepared by extensive dialysis against 10 mM sodium cacodylate-cacodylic acid at pH 6.1 or pH 6.8 containing 200 mM NaCl and 20 mM MgCl2.…”

“…EMSA experiments for the triplex formation were performed essentially as described previously by a 15% native polyacrylamide gel electrophoresis [12,13,[19][20][21][22][23][24][25][26][27]. In a 9 µl of reaction mixture, 32 P-labeled Pur23A•Pyr23T duplex (~1 nM) (Figure 1c) was mixed with increasing concentrations of the specific TFO (Pyr15TM, Pyr15NC7-1, Pyr15NC7-2, Pyr15NC5-1, or Pyr15NC5-2) (Figure 1c) and the nonspecific oligonucleotide (Pyr15NS2M) (Figure 1c) in buffer [50 mM Tris-acetate (pH 7.0), 100 mM NaCl, and 10 mM MgCl2].…”