CRISPR-Cas9 Genome Editing in Polyploid Oilseed Rape

In polyploid species, altering a trait by random mutagenesis is highly inefficient due to gene redundancy. The ALC gene is involved in valve-margin development and contributes to seed shattering from mature fruits. Knocking out ALC would increase shatter resistance and avoid seed loss during mechanical harvest. Janina Braatz from the Christian-Albrechts-University of Kiel in Germany, together with her team, used CRISPR-Cas9 to target two ALCATRAZ (ALC) homoeologs in tetraploid oilseed rape.

The team obtained a T1 plant with four alc mutant alleles. All mutations were stably inherited to the T2 progeny. The T2 generation was devoid of any wild type ALC alleles. T-DNA and ALC loci were not linked as indicated by random segregation in the T2 generation. Hence, double mutants lacking the T-DNA already could be selected in the first offspring generation.

While the whole genome sequencing data revealed at least five independent insertions of vector backbone sequences, the team did not detect any off-target effects in two genome regions homologous to the target sequence.

The simultaneous editing of multiple homoeologs in a polyploid species by CRISPR-Cas9 mutagenesis will offer new approaches for rapeseed breeding.

For more on this study, read the article in Plant Physiology.