Developing Efficient Genetic Transformation and Genome Editing Methods for Solanum americanum Mill., a Medicinal and Vegetable Plant

Abstract

Highly efficient tissue culture and transformation systems are indispensable for micropropagation, the molecular studies of secondary metabolism, the biotechnological enhancement of bioactive compounds, and genome editing in plants. Solanum americanum is a diploid species with both vegetable and medicinal value, yet remains largely underutilized. It produces a various pharmacologically active secondary metabolites, including glycoalkaloids, tropane alkaloids, and resveratrol. However, stable genetic transformation and reliable genome editing protocols for this species remain underdeveloped. In this study, we established an efficient genetic transformation and genome editing method for S. americanum. Our results showed that nearly all leaf explants responded positively to MS medium supplemented with 2 mg L−1 6-benzylamino purine (6-BA) and 0.1 mg L−1 α-naphthaleneacetic acid (NAA), generating an average of 25.85 adventitious shoots per explants within four weeks. Furthermore, among kanamycin-resistant shoots, approximately 89.7% exhibited normal growth and over 75% showed strong GFP expression as well as inheritance, which was confirmed through molecular analysis and fluorescence assays in flowers, fruits, and T1 progeny. Moreover, we successfully utilized this transformation method to achieve stable genome editing of the phytoene desaturase gene (SaPDS). Our results demonstrate that the tissue culture, transformation, and genome editing methods established for S. americanum provide a valuable toolkit for large-scale propagation, the study of secondary metabolism, and the biotechnological improvement of this species.