Application of an efficient adenine base editing system in developing Danshen (Salvia miltiorrhiza) dinitroaniline-resistant germplasm

Chinese herbal medicine Danshen (Salvia miltiorrhiza) has suggested therapeutic effects on cardiovascular and cerebrovascular diseases, inflammatory reactions and tumors, perhaps due to secondary metabolites, such as tanshinone. Danshen is sensitive to most broad-spectrum herbicides, thus it is a challenge to manage weeds during planting. Although CRISPR/Cas gene editing has been used to knock out important genes in Danshen, the more accurate base editing method has rarely been applied to this species. We developed an efficient adenine base editing system for Danshen and observed its editing effect in callus and transgenic plants. To demonstrate the efficacy of this technique, we generated several novel Danshen germplasms with mutant α-tubulins, some of which displayed resistance to a dinitroaniline herbicide, pendimethalin, in Danshen planting. We also found distinct effects of the different Danshen mutant tubulin homologues on dinitroaniline sensitivity, perhaps indicating a functional differentiation of the α-tubulin genes. Our work provides a new gene modification method for Danshen breeding, which might be extended to other medicinal plants.