Genome Editing Technologies for Resistance Against Phytopathogens: Principles, Applications and Future Prospects

Abstract

Genetic variation in crop plants is an indispensable factor for sustainable agriculture. For creating genetic variation, plant biotechnology relied on various random mutagenesis approaches such as γ-radiation, EMS generated mutagenesis etc. Now these methods are being replaced by genome editing technologies that precisely manipulate specific sequences in the genome. These technologies, based on sequence-specific nucleases (SSNs), mediated double-strand breaks in DNA at specific sites within the genome. Among them, three foundational targeted genome editing technologies are: TALENs (transcription activator-like effector nucleases), CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats) and ZFNs (zinc-finger nucleases). TALENs and ZNFs are synthetic engineered bipartite enzymes, consisting of two domains, (1) DNA binding domain and (2) Fok1 domain (nuclease).CRISPR/Cas 9 is a two-component unicellular machinery, with a sgRNA first component (which targets the specific sequence in genome), and Cas 9 as a second component (an enzyme that mediates site-specific targeting of genome). The Cas9 protein is DNA nuclease whose activity is determined by target seed sequence (first 20 nucleotides) of sgRNA. Therefore, multiple sgRNAs with different target seed sequences direct Cas9 to corresponding spots. This imperative characteristic of Cas9 enables it to edit at multiple sites simultaneously and imparts potential applications in both basic and applied research. By using these approaches, disease resistance in plants is achieved by knocking out those loci contributing in disease susceptibility and negative regulator genes in genome. Thereby these techniques are becoming a new toolbox of every modern molecular biology laboratory for editing plant genome. This chapter provides a detailed overview of genome editing approaches for developing disease resistant in plants, describing comprehensive knowledge on genome editing principles, application of these techniques, challenges and prospects.