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Mendel-Brno 2000

category image Volume: 17
Issue Number 6, Part 2
June 2000

Padlock Oligoucleotides for Duplex DNA based on sequence specific triple helix formation

Triple helix forming oligonucleotides can bind in the major groove of double-stranded DNA by establishing non-covalent interactions. We have developed a new method which brings the strength of the covalent bond to interactions between an oligonucleotide and a plasmid, without any enzymatic or chemical modification of the target double-stranded DNA, through the use of a triple-helical complex intermediate. After sequence specific recognition of a double-stranded DNA target through triple helix formation, the ends of the triplex-forming oligonucleotide were joined through the action of T4 DNA ligase, thus creating a circular DNA molecule catenated to the plasmid containing the target sequence. Different types of triple helices can be used in this method. We have shown using a restriction enzyme inhibition assay that the circular oligonucleotide is free to move along the plasmid under conditions where triple helix is not stable. These padlock oligonucleotides provide a new tool to attach in an irreversible way a non-covalent tag to supercoiled plasmids or other double-stranded DNAs. They could be used for example for the labeling or purification of plasmids, or in targeted plasmid delivery. Recent developments in this field will be presented.

References

Escude, C.,Garestier, T. & Helene, C. (1999). Padlock Oligonucleotides for Duplex DNA Based on Sequence Specific Triple Helix Formation.Proc. Natl. Acad. Sci. U S A 96, 10603-10607.

Christophe Escude, Thibaut Roulon, Therese Garestier and Claude Helene.

Laboratoire de Biophysique, Museum National d'Histoire Naturelle,
INSERM U 201, CNRS UMR 4686,
43, rue Cuvier, 75231 Paris Cedex 05, France. ,
email: escude@mnhn.fr

$15.00