Somatic embryogenesis research facilitates citrus improvement via cell engineering

Somatic embryogenesis (SE) is a key regeneration pathway in various cell engineering approaches to crop improvement, especially for economically important perennial woody crops like citrus.
However, SE capability is difficult to maintain, and recalcitrance to SE has become a major obstacle to plant in vitro breeding.
To understand the mechanism of SE in citrus, we profiled 50 known and 45 novel miRNAs and their target genes in the nonembryogenic callus (NEC) and embryogenic callus (EC). The conserved csi-miR156a/b and csi-miR171c were preferentially accumulated in the embryogenic callus.
Overexpression of csi-miR156a and RNAi-mediated suppression of individual SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (CsSPL) target genes promote SE. miR156a-CsSPLs modulate starch accumulation in EC cells through miR172d-CsTOEs, and thus regulate SE. Likewise, overexpression of miR171 and suppression of SCARECROW-LIKE2 (CsSCL2) target genes promote SE. CsSCL2/3 modulate reactive oxygen species (ROS) homeostasis through the interactive protein CsClot of the thioredoxin superfamily and directly suppress the expression of regeneration-related genes, and thus regulate SE. In citrus, the protoplasts isolated from EC are capable of regeneration through SE approach.
We optimized citrus protoplast transfection system to an efficiency of 68% and achieved gene editing in the EC protoplasts with an efficiency over 10%, which provides a promising system for regenerating transgene-free but gene-edited citrus plants.

XV International Citrus Congress

Xiao-Meng Wu, Meng-Qi Feng, Wen-Hui Yang, Ru-Meng Quan, Wen-Wu Guo

citrus, embryogenic callus, somatic embryogenesis, miR156-CsSPL, miR171-CsSCL, protoplast transfection

https://doi.org/10.17660/ActaHortic.2026.1448.31