Establishment and Validation of an Efficient Agrobacterium Tumefaciens-Mediated Transient Transformation System for Salix Psammophila

<i>Salix psammophila</i>, C. Wang & Chang Y. Yang, a desert-adapted shrub, is recognized for its exceptional drought tolerance and plays a vital role in ecosystem maintenance. However, research on <i>S. psammophila</i> has been limited due to the lack of an efficient and reliable genetic transformation method, including gene functional studies. The <i>Agrobacterium</i>-mediated transient overexpression assay is a rapid and powerful tool for analyzing gene function in plant vivo. In this study, tissue culture seedlings of <i>S. psammophila</i> were utilized as the recipient materials, and the plant expression vector pCAMBIA1301, containing the <i>GUS</i> reporter gene, was transferred into the seedlings via an <i>Agrobacterium</i>-mediated method. To enhance the efficiency of the system, the effects of secondary culture time, <i>Agrobacterium</i> concentration, infection time, and co-culture duration on the transient transformation efficiency of <i>S. psammophila</i> were explored. The optimal combination for the instantaneous transformation of <i>S. psammophila</i> tissue culture seedlings mediated by <i>Agrobacterium</i> was determined as follows: a secondary culture time of 30 d, a value of OD₆₀₀ of 0.8, an infection time of 3 h, and a co-culture duration of 48 h. Subsequently, the effectiveness of the transformation system was validated using the <i>S. psammophila</i> drought response gene <i>SpPP2C80</i>. To further confirm the accuracy of the system, <i>SpPP2C80</i>-overexpressing <i>Arabidopsis</i> was constructed and drought resistance analysis was performed. The results were consistent with the transient overexpression of <i>SpPP2C80</i> in <i>S. psammophila</i> tissue culture seedlings, indicating that this system can be effectively employed for studying gene function in <i>S. psammophila</i>. These findings provide essential information for investigating gene function in non-model plants and pave the way for advancements in molecular biology research in <i>S. psammophila</i>.