CRISPR/Cas9-mediated gene editing tool and fathomless genetic and metabolic engineering applications in plants

CRISPR-/Cas9-mediated genome editing is modernizing plant science by presenting simple, accurate and high-throughput tools for genetic engineering in various plant species. This modern technology utilizes guide RNAs, which direct Cas9 endonuclease, to generate double-stranded breaks at targeted sites of the genome, and finally plant DNA repair by either nonhomologous end-joining or homology-directed repair mechanism resulted in an error-prone efficient mutation in the genome. However, the targeted modification of the plant genome depends upon the selection of target sites, vector carrying its important components and its delivery method in plant cells. In this chapter, we described different types of CRISPR/Cas9 plant transformation vectors, its properties and delivery methods in plant cells. Initially, the cloning vectors harbouring the single-guide RNA and Cas9 gene regulated by different promoters were used for genome editing in plant protoplast. Further, binary Ti plasmid carrying T-DNA and broad host-range replicon was used for the development of different CRISPR/Cas9 plant transformation vectors. These CRISPR/Cas9 vectors carrying essential components are delivered to plant cells via PEG-mediated protoplast transfection, A. tumefaciensmediated transformation or transformation by bombardment method.