Talen- and CRISPR/Cas9-Based Gene Editing in Maize and Rice

Modified CRISPR (clustered regularly interspaced short palindromic repeats) systems, comprising Cas9 endonucleases and single guide RNAs (gRNAs), and TALENs (TAL effector nucleases) have emerged as potent biotechnological tools for both basic and applied research. The most promising utilization of both Cas9/sgRNA and TALENs is for targeted genome editing, precise genetic alterations within any genome of interest, as demonstrated in a plethora of organisms including several crop plants. My presentation describes development and application of these two technologies to generate heritable genome modifications in rice and maize. TALENs were employed to generate stable, heritable mutations in dozens of rice genes and maize glossy2 gene. The frequency of mutagenesis varied from species and constructs targeting different genes. The highest rate in rice reached about 65% of transgenic lines carry desired mutations, while construct targeting the maize glossy2 gene produced mutant lines from the maize genotype Hi-II at a frequency of about 10%. Phenotypic changes associated with mutated genes were observed in some mutant lines in rice and maize. We also modified the Cas9/sgRNA system suitable for targeted gene mutagenesis in rice and maize, respectively. The two systems have been successfully applied to rice and maize for targeted mutagenesis of many genes. Transgenic lines of T0 generation carrying site-specific mutations were produced at frequency as high as 100% in rice and 80% in maize. Our results demonstrate that TALENs and Cas9/sgRNA are effective toolboxes for genome editing in rice and maize, empowering the discovery of gene function and the trait improvement.