A rapid and efficient transient expression system for gene function and subcellular localization studies in the tea plant (Camellia sinensis) leaves

Tea, the most consumed non-alcohol beverage in the world, has many specialized secondary metabolites, and functional identification of their related genes is very important for the improvement of tea quality. However, a transient transformation system, which is considered as a valuable tool for gene function and protein subcellular localization, has not been established in tea plants. Here, we report a rapid and efficient system for transient transformation of tea plant leaves using antisense oligodeoxynucleotide (asODN) for gene suppression and Agrobacterium for gene over-expression. Specifically, we established the transient transformation system by analyzing different varieties, injection solution concentrations, and sampling time post injection, followed by green fluorescent protein (GFP) fluorescence signal and western blot analysis to confirm the reliability of the over-expression transient transformation system. Additionally, the suppression and over-expression effects of important genes relate to secondary metabolism were evaluated at gene expression and metabolic levels. Finally, a transcription factor was transiently over-expressed to determine its subcellular location by protoplasts successfully in the tea plant leaves. Our data indicate that this system is reliable and effective for studying gene function and protein localization in tea plant leaves, providing new insights into tea plant biology research.