CRISPR-Cas9 has proved to be a powerful genome-editing tool for introducing genetic changes into crop species. However, it has not yet been used to edit cassava (Manihot esculenta). To test the capacity of CRISPR-Cas9 in cassava, Donald Danforth Plant Science Center researcher John Odipio and his team targeted the phytoene desaturase (MePDS) gene in two cultivars using constructs carrying gRNAs targeting MePDS. Modification of the MePDS gene generates visually-detectable mutated events in a relatively short time frame and does not require sequencing to confirm.
After Agrobacterium-mediated delivery of CRISPR-Cas9 reagents into cassava cells, both constructs induced visible albino phenotypes within cotyledon-stage somatic embryos and plants were regenerated from there. A total of 38 lines, 19 from each cultivar, were analyzed for mutagenesis. The frequency of plant lines showing albino phenotype ranged from 90 to 100% for both cultivars. Sequence analysis revealed that all the lines examined carried mutations at the targeted MePDS site, with insertions, deletions, and substitutions recorded. The team observed minor nucleotide substitutions and or deletions upstream of the 5′ end or downstream of the 3′ targeted MePDS region.
The data reported in this study demonstrates that CRISPR-Cas9-mediated genome editing also applies to cassava, with high efficiency. Read more