To assess germinal transmission of GEX1A resistance among SGR mutants, Butt and colleagues carried out genetic and phenotypic analysis in the next generation. Homozygous mutants were phenotypically indistinguishable from the wild-type plants, suggesting that these SF3B1 variants exhibit full splicing activity in rice. The resistance to GEX1A, however, is dose dependent and variable among SGR mutants. The mutant SGR4 displayed the strongest resistance to GEX1A. The seeds of this mutant can establish well on medium with GEX1A as high as 10 μM; under the same conditions other SGR mutants failed to germinate. Although SGR4 carried three mutations, it is likely the K1050E missense mutation has largely contributed to weakening the SF3B1 and GEX1A interaction.

This study demonstrated that it is feasible to conduct directed evolution in plants, which has significant implications. Plants evolve to adapt to their growth environments in a typically lengthy process. Accelerated evolution may provide an efficient pathway to achieving high agriculture productivity and food security in the face of global warming and climate change. Given the enormous sizes of crop genomes, it is effectively impossible to achieve saturating mutagenesis in vivo. With CRISPR, near-saturation mutagenesis becomes achievable, as shown in this study. Hence, such a directed evolution approach will be very powerful for evolving and engineering beneficial traits in crops such as herbicide resistance, improved photosynthesis, and enhanced tolerance or resistance to abiotic or biotic stresses. Genomics tools such as genome-wide association studies (GWASs) have aided rapid mapping and discovery of important genes and alleles in crop productivity and stress tolerance in major crops. It is anticipated that CRISPR-enabled directed evolution on these target genes would help improve agronomic traits or generate novel alleles directly in elite cultivars. While directed evolution is often used for protein engineering, it is also possible to apply it towards engineering quantitative traits by targeting cis-regulatory elements [7]. Furthermore, we envision that similar directed evolution approaches can be practiced in animals as well, such as worm, fruit fly, and zebrafish.