New Toolkit for Plant Genome Engineering

The Science

Develop plant genome engineering techniques to allow efficient gene knockouts, replacements, altered transcriptional regulation, and multiplexed modifications.

The Impact

• The toolkit was tested for deletion mutagenesis targeting single or multiple genes, taking advantage of polycistronic transcripts with multiple guide RNAs (gRNA) in a single construct.
• Up to 6 genes were successfully knocked out using multiplexed vectors, and a 58 Kbp genomic region containing 5 genes involved in nitrogen fixation was deleted in a single event.
• The new toolkit will accelerate genome engineering of bioenergy crops, providing a flexible and comprehensive set of gene-editing options.

Summary

• A suite of direct and modular cloning vectors was developed to build TALEN-or CRISPR/Cas9-based editing targeting multiple genes and containing different promoters, reporters, selectable markers, and codon-optimized genes.
• Vector construction is facilitated by an online tool that allows choice of different vector modules for different functions, either in monocot or dicot plants.

References

Cermák, T. et al. “Multipurpose toolkit to enable advanced genome engineering in plants.” The Plant Cell 29(6), 1196–1217 (2017). [DOI:10.1105/tpc.16.00922].