Haixia Institute of Science and Technology, State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops, College of Life Sciences, College of Forestry, Fujian Agriculture and Forestry University, Fuzhou, 350002, China.
Planta. 2024 Jun 5;260(1):18. doi: 10.1007/s00425-024-04448-5.
We have developed and optimized a rapid, versatile Agrobacterium-mediated transient expression system for cannabis seedlings that can be used in functional genomics studies of both hemp-type and drug-type cannabis. Cannabis (Cannabis sativa L.) holds great promise in the medical and food industries due to its diverse chemical composition, including specialized cannabinoids. However, the study of key genes involved in various biological processes, including secondary metabolite biosynthesis, has been hampered by the lack of efficient in vivo functional analysis methods. Here, we present a novel, short-cycle, high-efficiency transformation method for cannabis seedlings using Agrobacterium tumefaciens. We used the RUBY reporter system to monitor transformation results without the need for chemical treatments or specialized equipment. Four strains of A. tumefaciens (GV3101, EHA105, LBA4404, and AGL1) were evaluated for transformation efficiency, with LBA4404 and AGL1 showing superior performance. The versatility of the system was further demonstrated by successful transformation with GFP and GUS reporter genes. In addition, syringe infiltration was explored as an alternative to vacuum infiltration, offering simplicity and efficiency for high-throughput applications. Our method allows rapid and efficient in vivo transformation of cannabis seedlings, facilitating large-scale protein expression and high-throughput characterization studies.
我们开发并优化了一种快速、通用的农杆菌介导的大麻幼苗瞬时表达系统，可用于大麻的工业大麻和毒品大麻类型的功能基因组学研究。大麻（Cannabis sativa L.）由于其多样化的化学成分，包括特殊的大麻素，在医学和食品工业中具有巨大的应用前景。然而，涉及各种生物过程的关键基因的研究，包括次生代谢物生物合成，由于缺乏有效的体内功能分析方法而受到阻碍。在这里，我们提出了一种使用根癌农杆菌的新型、短周期、高效的大麻幼苗转化方法。我们使用 RUBY 报告系统来监测转化结果，而无需进行化学处理或使用特殊设备。我们评估了四种根癌农杆菌（GV3101、EHA105、LBA4404 和 AGL1）的转化效率，发现 LBA4404 和 AGL1 的性能更优。该系统的多功能性还通过 GFP 和 GUS 报告基因的成功转化得到了进一步证明。此外，我们还探索了注射器浸润作为真空浸润的替代方法，为高通量应用提供了简单高效的方法。我们的方法允许快速有效地对大麻幼苗进行体内转化，促进了大规模的蛋白质表达和高通量的表征研究。
