Efficient isolation of protoplasts from freesia callus and its application in transient expression assays
2019
Shan, Xiaotong | Li, Yueqing | Zhou, Liudi | Tong, Linna | Wei, Chao | Qiu, Lijun | Gao, Xiang | Wang, Li
Freesia hybrida is one of the most famous cut flower variety domesticated from wild species in Freesia genus, distinguished for its colorful flower colors and sweet scents. It has potential to be a model plant to unravel the molecular basis of plant specialized metabolites biosynthesis in family Iridaceae, which are crucial to genetically modify ornamental traits such as flower color and flower scents. However, the lengthy life cycle propagated from tissue culture and extremely low transformation rate severely decelerate functional characterization of genes through transgenic method in this iridaceous plant. Using calluses induced from young flower buds from one popular cultivar of Freesia hybrida, ‘Red River®’, an efficient protoplast isolation method was established and optimized, which was further employed in the gene transient expression system. Results demonstrated that the newly explored transient system was versatile and could be applied in numerous aspects, such as protein immunoblotting, enzyme activity, protein subcellular localization, protein–protein interaction, protein–DNA interaction and regulatory gene transregulation property assays. Moreover, calluses induced from another cultivar of Freesia hybrida, ‘Ambiance’ could also be used to isolate protoplasts for gene transient transfection analysis, indicating the wide applicability of this system. Results in this study provided new methods for the elucidation of gene function in Freesia in vivo, which might also be inspirational in other species with long life cycles or hardly to be transformed, such as plants in family Iridaceae.
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