Establishment of an Agrobacterium-Mediated Genetic Transformation System for Prunus sibirica and Its Preliminary...

Prunus sibirica possesses substantial ecological and economic value and is recognized as an emerging woody energy crop; however, its regeneration and genetic transformation remain challenging. The present study established an efficient regeneration and Agrobacterium tumefaciens-mediated transformation system for P. sibirica. Stem segments served as explants, and the optimal proliferation medium was Murashige and Skoog medium supplemented with 0.8 mg L^-1 6-benzylaminopurine (6-BA), 0.2 mg L^-1 thidiazuron (TDZ), and 0.1 mg L^-1 1-naphthaleneacetic acid (NAA), resulting in a proliferation coefficient of 6.53. The genetic transformation system utilized A. tumefaciens strain GV3101 carrying reporter vectors pRI101-GFP or pCAMBIA1301-GUS, with optimal parameters including a 3-day preculture, infection at OD₆₀₀ = 0.6 for 20 min, and a 2-day coculture. Transformants were selected in a medium containing 20 mg L^-1 kanamycin and 300 mg L^-1 cefotaxime, achieving an average transformation efficiency of 11.27%. As an initial application, PsWRKY17-overexpressing lines were generated, exhibiting transcript levels 7-18 times higher than those in the empty-vector control. Following low-temperature treatment, PsWRKY17-OE lines demonstrated increased SOD and POD activities and reduced relative electrical conductivity and MDA content compared to the control (p < 0.01). These results provide preliminary physiological evidence that PsWRKY17 overexpression is associated with altered low-temperature stress responses in P. sibirica. However, the regulatory mechanism of PsWRKY17 requires further investigation. In summary, an effective regeneration and genetic transformation system for P. sibirica was developed, providing a foundation for future research and biotechnological applications.