CRISPR/CAS GENOME EDITING TECHNOLOGY FOR CROP IMPROVMENT

The accelerated growth in the world population together with the process of global warming call for breeding of crops capable of improved yielding under extreme climate conditions and with enhanced tolerance to biotic stresses. Mutants with improved stress tolerance have been identified in model plants. However, the lack of an efficient plant gene knockout technology hindered the development of similar stress-resistant crops. Recently, a flexible targeted genome editing technology based on a bacterial clustered, regularly interspaced, short palindromic repeat (CRISPR)-associated protein 9 nuclease from Streptococcus pyogenes (Cas9) has been developed. In this system, Cas9 bound to a synthetic guide RNA can induce sequence specific genomic DNA double-strand break (DSB). DSBs are repaired mainly by the erroneous non-homologous end joining system which leads to loss of bases around the break. Occasionally, this results in the formation of premature stop codon and gene knockout. Thus, the CRISPR-Cas9 is a promising technology that can knockout targeted genes without rendering corresponding plants as classical GMO, paving the way to its implementation in agricultural biotechnology. We have utilized this technology to develop novel crops with improved biotic and abiotic stress-resistance. These include extreme temperatures tolerant tomato and virus resistant cucumber cultivars. In addition, the CRISPR-Cas9 technology was utilized to knockout genes involved in the maintenance of heritable methylation in vegetable crops. This is expected to expose stable epi-phenotypes that will serve as a unique resource to identify beneficial epiallels in the areas of stress resistance. The generation and characterization of the different CRISPR-Cas9 modified crops will be discussed.