Genome Editing in Apple

Genome editing with artificially engineered nucleases is an advanced molecular technology for pursuing precise and effective genetic engineering. In this technology, engineered nucleases induce DNA double-strand breaks at targeted sites in a genome, stimulating the DNA repair system in cells, thus enabling site-directed mutagenesis. Genome editing using CRISPR (clustered regularly interspaced short palindromic repeats)/CRISPR-associated protein9 (Cas9), originating from a defence system of prokaryotes, is a powerful technology that is now being widely utilized in molecular research studies, as well as in breeding programmes of various plant species, including fruit trees, to impart either novel or enhanced traits to established commercial cultivars or to new cultivars/genotypes. Recently, several reports have demonstrated successful apple genome editing and the introduction of important traits, such as those for early flowering and reduced fire blight susceptibility, to popular commercial cultivars, such as ‘Gala’ and ‘Golden Delicious’. It is important to point out that these reports reveal that such genome-edited/mutant apple plants or cell lines do not carry foreign genes. Nevertheless, during the process of precise genome editing, the coexistence of various types of mutations referred to as “mosaic mutations” and off-target effects are major concerns. Therefore, to minimize such effects, selection of target sequences and estimation of off-target effects for CRISPR/Cas9 has been developed for many organisms, and these have also been employed for apple by using in silico analysis based on genome information. On the other hand, apple genome heterozygosity has led to difficulties in genome editing, as the complex genome of apple precludes the use of some of these basic techniques for genome editing. Therefore, further studies focused on genome information and culture techniques tailored for apple are needed. It will be highly critical for each apple cultivar in developing precise and efficient genome editing for apple. This chapter will provide an overview of current studies of genome editing in apple and will discern and explore how this strategy will provide insights into molecular breeding technologies for genetic improvement of the apple.