Artificial Restriction DNA Cutters to Promote Homologous Recombination in Human Cells

Hitoshi      Katada; Makoto      Komiyama

doi:10.2174/156652311794520094

Abstract

Homologous recombination is almost the only way to modify the genome in a predetermined fashion, despite its quite low frequency in mammalian cells. It has been already reported that the frequency of this biological process can be notably increased by inducing a double strand break (DSB) at target site. This article presents completely chemistrybased artificial restriction DNA cutter (ARCUT) for the promotion of homologous recombination in human cells. This cutter is composed of Ce(IV)/EDTA complex (molecular scissors) and two strands of peptide nucleic acid (PNA), and contains no proteins. Its scission site in the genome is determined simply by Watson-Crick rule so that ARCUT for desired homologous recombination is easily and straightforwardly designed and synthesized. The site-specificity of the scission is high enough to cut human genome at one target site. The DSB induced by this cutter is satisfactorily recognized by the repair system in human cells and promotes the targeted homologous recombination.

Keywords: Genome manipulation, artificial restriction DNA cutter, cerium, peptide nucleic acid, homologous recombination, double strand break, phosphodiester, thymine, pcPNA strands, hypermethylation, X syndrome, fluorescent protein, green fluorescence, adenovirus vector