Gene editing of three BnITPK genes in tetraploid oilseed rape leads to significant reduction of phytic acid in seeds.

Commercialization of Brassica napus. L (oilseed rape) meal as protein diet is gaining more attention due to its well-balanced amino acid and protein contents. Phytic acid (PA) is a major source of phosphorus in plants but is considered as anti-nutritive for monogastric animals including humans due to its adverse effects on essential mineral absorption. The undigested PA causes eutrophication, which potentially threatens aquatic life. PA accounts to 2-5% in mature seeds of oilseed rape and is synthesized by complex pathways involving multiple enzymes. Breeding polyploids for recessive traits is challenging as gene functions are encoded by several paralogs. Gene redundancy often requires to knock out several gene copies to study their underlying effects. Therefore, we adopted CRISPR-Cas9 mutagenesis to knock out three functional paralogs of BnITPK. We obtained low PA mutants with an increase of free phosphorus in the canola grade spring cultivar Haydn. These mutants could mark an important milestone in rapeseed breeding with an increase in protein value and no adverse effects on oil contents.