Gene Targeting by Homologous Recombination

Abstract

Abstract Gene targeting by homologous recombination involves the exchange of genetic information between genomic and exogenous deoxyribonucleic acid (DNA) molecules via crossover events. These exchanges are guided by homologous sequences acted on by enzymatic machinery of the cell. Homologous recombination provides a mechanism for targeting defined modifications into genes of interest, making gene‐targeting technologies valuable tools to explore gene function and to develop human, genetic disease models. Gene targeting, however, is inefficient, making the process challenging. Advances in technology now allow us to direct repair enzymes to the targeted site by inducing site‐specific DNA damage using zinc‐finger nucleases, transcription activator‐like effector nucleases, clustered regularly interspaced short palindromic repeats and triplex‐forming oligonucleotides. Further, antirecombinogenic pathways can now be transiently suppressed using ribonucleic acid (RNA) interference (RNAi) and other small molecule approaches. These and other techniques can greatly enhance gene‐targeting efficiency. The coincident development of human stem cell technology brings forth the potential of gene‐targeting strategies for therapeutic application. Key Concepts: Homology‐directed gene targeting utilises homologous recombination to introduce defined modifications into sequences of interest in mammalian genomes. Gene‐targeted knockout and knock‐in models are instrumental in elucidating gene function and studying human genetic diseases. Combining stem cell and gene‐targeting technologies opens potential avenues for gene therapy. Recent technological advances have greatly enhanced gene‐targeting efficiencies. Due to technological advances, most genes in a variety of mammalian species can now be manipulated by homology‐directed gene targeting.