Targeted gene modification by oligonucleotides and small DNA fragments in eukaryotes.
Targeted modification of genomic DNA (gDNA) in a cell permanently changes its phenotype. Numerous methods to modify target portions of gDNA in eukaryotes have been developed, and several have found practical application (e.g., homologous recombination-utilized modification in mouse ES cells). The past decade has seen active research both in oligonucleotide-directed modifications (where the oligonucleotides can be chimeric RNA/DNA oligonucleotides, single-stranded oligonucleotides, or triple helix-forming oligonucleotides), and in small DNA fragment-directed targeted modifications. These modifications can alter gDNA at several basepairs. In order to specifically direct the reaction to the targeted site, short oligonucleotides (shorter than 80-mers) or small DNA fragments (500 to approximately 2000 bps) are needed. Such oligonucleotides and small DNA fragments can be prepared easily. Although the mechanism by which targeted modification occurs has not been fully elucidated, numerous eukaryotic target genes have been modified using these methods in both cultured cells and live animals. The effectiveness of each method has been confirmed, but the efficiency of modification for all these approaches is presently insufficient for practical use. Further studies, including elucidation of complete mechanisms and optimization of methods for introducing oligonucleotides or small DNA fragment into cell nuclei, are required.
Suzuki
Takayuki
T
Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya 464-8601, Japan. tsuzuki@nagoya-u.jp
D000818:Animals; D004247:DNA / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000737:chemistry*; D053595:Embryonic Stem Cells / Q000166:cytology; D018014:Gene Transfer Techniques; D005821:Genetic Techniques; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008957:Models, Genetic; D009841:Oligonucleotides / Q000737:chemistry*; D011995:Recombination, Genetic
D004277:DNA, Single-Stranded; D009841:Oligonucleotides; D004247:DNA