Gene targeting by triple helix-forming oligonucleotides.
Effective gene targeting reagents would have widespread utility for genomic manipulation including transgenic cell and animal construction and for gene therapy. They would also be useful in basic research as probes of chromatin structure, and as tools for studying the repair and mutagenesis of targeted DNA damage. We are developing triple helix-forming oligonucleotides (TFOs) for gene targeting in living mammalian cells. Challenges to TFO bioactivity include the impediments to the biochemistry of triplex formation presented by the physiological environment and the charge repulsion between the duplex and the third strand. In addition, there are biological constraints to target access imposed by mammalian chromatin structure. Here we describe the oligonucleotide modification format that appears to support biological activity of TFOs. In addition we show that manipulation of the cell biology, specifically the cell cycle, has a dramatic influence on TFO bioactivity.
Majumdar
Alokes
A
Laboratory of Molecular Gerontology, National Institute on Aging, National Institutes of Health, Baltimore, Maryland 21224, USA.
Puri
Nitin
N
McCollum
Nicholas
N
Richards
Sally
S
Cuenoud
Bernard
B
Miller
Paul
P
Seidman
Michael M
MM
D000818:Animals; D002453:Cell Cycle / Q000502:physiology; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000187:drug effects*; D018390:Gene Targeting; D005819:Genetic Markers; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000494:pharmacology*
D003432:Cross-Linking Reagents; D005819:Genetic Markers; D009838:Oligodeoxyribonucleotides; D004247:DNA; D007041:Hypoxanthine Phosphoribosyltransferase