Targeted mutagenesis and functional marker development of two Bna.TAC1s conferring novel rapeseed germplasm with compact architecture.

Abstract

Key message: Simultaneous disruption of two Bna.TAC1s, redundantly controlling the branch angle, generates a compact architecture in rapeseed, and two functional markers are developed to facilitate breeding rapeseed cultivars with compact architecture. Shoot branch angle is a key factor in determining the aerial plant architecture. A narrow branch angle can increase yields by facilitating mechanized harvest and high-density planting in rapeseed, a globally important oil crop. However, the available rapeseed varieties with narrow branch angle are very limited. In this study, two Bna.TAC1 members named BnaA5.TAC1 and BnaC4.TAC1 were found to have the four canonical domains of TAC1-like members, including domains I, II, III and IV in rapeseed. Each Bna.TAC1 exhibits dominant expression in the lateral branch with gradual dynamic response to light and encodes a protein localized in the plasma membrane. CRISPR/Cas9-mediated editing system was used to simultaneously knock out the two Bna.TAC1s to obtain two different Bna.tac1 double mutants, designed as CR-Bna.tac1-1 and CR-Bna.tac1-2. These two mutants displayed different degrees of compact architecture without affecting plant height and yield-related traits. The two Bna.TAC1s were also shown to play a redundant role in controlling branch angle by regulating the gravitropic response. In addition, we developed two specific gel-based functional markers in each Bna.TAC1 for the transgene-free mutant CR-Bna.tac1-1, which co-segregate with narrower branch angle and could help to identify the mutant alleles in a segregating population. We also found that the genomic variation of the two Bna.TAC1s is not associated with branch angle variation in the natural rapeseed population. Overall, these results reveal the key roles of Bna.TAC1s in regulation of rapeseed branch angle and provide a novel germplasm and functional markers for breeding superior varieties with compact architecture in rapeseed.