Genome Editing Strategies for Rice Improvement.

The precision and ease of use of CRISPR nucleases, such as Cas9 and Cpf1, for plant genome editing has the potential to accelerate a wide range of applications for crop improvement. For upstream research on gene discovery and validation, rapid gene knock-outs can enable testing of single genes and multi-gene families for functional effects. Large chromosomal deletions can assist with positional cloning of QTLs by helping to narrow down the target region. Nuclease-deactivated Cas9 fusion proteins with transcriptional activators and repressors can be used to up and down-regulate gene expression. Even more promising, gene insertions and allele replacements can provide the opportunity for rapidly testing the effects of differing alleles at key loci in the same genetic background. The rice community is well positioned to take advantage of these new opportunities, as the wealth of cloned genes, diverse genetic donors, and sequence data present numerous testable hypotheses with clear implications for rice improvement. For example, data from the 3,000 Rice Genomes Project and High Density Rice Array (HDRA) studies can be used to define sequence variation at candidate genes underlying GWAS hits, which can then be rapidly validated using CRISPR editing techniques. Likewise, beneficial genes and QTLs originally identified from exotic germplasm, including landraces and wild relatives, can be rapidly transferred into elite breeding materials without causing negative linkage drag. Towards this end, a survey was performed to identify promising genes that can be used in a genome editing pipeline for rice improvement. We are currently exploring two approaches to develop efficient tools for rice breeding: (1) using CRISPR/Cas9 genome editing of elite rice cultivars to modify specific traits; and (2) editing exotic landraces having valuable traits to remove negative characters that prevent them from being used as efficient genetic donors in breeding programs.