Victoria Roig

Solid-phase preparation of 5′,3′-heterobifunctional oligodeoxyribonucleotides using modified solid supports

Abstract The solid-phase preparation of oligodeoxyribonucleotides attached to intercalator or reactive groups through their 5′- and (or) 3′-ends is reported. These syntheses implicate the introduction of suitable masked functional groups at the 5′-end of the oligonucleotide by the intermediate of their phosphoramidite derivatives or at the 3′-end of the oligonucleotide using modified solid supports. After full deblocking, the functional groups (phosphate, thiophosphate, primary amine or thiol) can be reacted with the suitable reactive group involved in the chosen ligand. These methods allow the preparation of heterobifunctional derivatized oligodeoxyribonucleotides.

Triplex-forming oligonucleotide–orthophenanthroline conjugates for efficient targeted genome modification

The inefficiency of gene modification by homologous recombination can be overcome by the introduction of a double-strand break (DSB) in the target. Engineering the endonucleases needed, however, remains a challenging task that limits widespread application of nuclease-driven gene modification. We report here that conjugates of orthophenanthroline (OP), a DNA cleaving molecule, and triplex-forming oligonucleotides (TFOs), known to bind specific DNA sequences, are synthetic nucleases efficient at stimulating targeted genome modification. We show that in cultured cells, OP-TFO conjugates induce targeted DSBs. An OP-TFO with a unique target was highly efficient, and mutations at the target site were found in ≈10% of treated cells, including small deletions most likely introduced during DSB repair by nonhomologous end joining. Importantly, we found that when homologous donor DNA was cotransfected, targeted gene modification took place in >1.5% of treated cells. Because triplex-forming sequences are frequent in human and mouse genes, OP-TFO conjugates therefore constitute an important class of site-specific nucleases for targeted gene modification. Harnessing DNA-damaging molecules to predetermined genomic sites, as achieved here, should also provide inroads into mechanisms of DNA repair and cancer.

Oligo-α-deoxyribonucleotides with a modified nucleic base and covalently linked to reactive agents

Abstract Solid-phase preparation of oligo-α-deoxyribonucleotides attached to intercalator, chemically or photochemically reactive groups through either their 5′- or 3′-ends, including use of the 5-methyl-α-deoxycytidine.

Synthesis and characterization of o6-modified deoxyguanosine-containing oligodeoxyribonucleotides for triple-helix formation

Abstract Two new modified deoxyguanosine derivatives linked through their C-6 position to a psoralen and an acridine derivative have been synthesized and incorporated into oligonucleotide chains. Difficulties observed during the purification of oligonucleotides containing a stretch of G and one method used to solve this problem are discussed.

Oligo-β- and -α-deoxyribonucleotides involving 2-aminopurine and guanine for triple-helix formation

Abstract Purine oligo-β-deoxyribonucleotides and unnatural nuclease-resistant α-anomer derivatives were constructed with 2-aminopurine (amP) guanine (G) in order to recognize purine sequences in double-helical DNA via isomorphous base triples amP.AT and G.GC.