Genome Editing as a Tool for Fruit Ripening Manipulation

Over the last few years, a series of tools for genome editing have been developed, allowing the introduction of precise changes into plant genomes. These have included Zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and CRISPR/Cas9, which is so far the most successful and commonly used approach for targeted and stable editing of DNA, due to its ease of use and low cost. CRISPR/Cas9 is now being widely used as a new plant breeding technique to improve commercially relevant crop species. Fruit ripening is a complex and genetically controlled developmental process that is essential for acquiring quality attributes of the fruit. Although the number of studies published to date using genome editing tools to molecularly understand or improve fruit ripening is scarce, in this review we discuss these achievements and how genome editing opens tremendous possibilities not only for functional studies of genes involved in fruit ripening, but also to generate non-transgenic plants with an improved fruit quality

Targeted Inter-Homologs Recombination in Arabidopsis Euchromatin and Heterochromatin

Homologous recombination (HR) typically occurs during meiosis between homologs, at a few unplanned locations along the chromosomes. In this study, we tested whether targeted recombination between homologous chromosomes can be achieved via Clustered Regulatory Interspaced Short Palindromic Repeat associated protein Cas9 (CRISPR-Cas9)-induced DNA double-strand break (DSB) repair in Arabidopsis thaliana. Our experimental system includes targets for DSB induction in euchromatic and heterochromatic genomic regions of hybrid F1 plants, in one or both parental chromosomes, using phenotypic and molecular markers to measure Non-Homologous End Joining and HR repair. We present a series of evidence showing that targeted DSBs can be repaired via HR using a homologous chromosome as the template in various chromatin contexts including in pericentric regions. Targeted crossover was rare, but gene conversion events were the most frequent outcome of HR and were found in both “hot and cold” regions. The length of the conversion tracts was variable, ranging from 5 to 7505 bp. In addition, a typical feature of these tracks was that they often were interrupted. Our findings pave the way for the use of targeted gene-conversion for precise breeding