Agrobacterium -mediated gene delivery and transient GFP expression in taro embryogenic cell suspensions

Embryogenic suspension cultures (ECS) of taro cultivars CPUK and THA-07 were subjected to genetic transformation using Agrobacterium tumefaciens strain AGL1 carrying the plasmid pART-TEST7, which included the neomycin phosphotransferase II ( npt II) selectable marker gene and the green fluorescent protein (gfp) as a reporter gene. The ECS were co-cultivated with the bacterial suspension in darkness at 23 °C for three days. Transformation efficiency was affected by several factors: Agrobacterium density, acetosyringone concentration, type of co-cultivation medium, temperature, genotype, and whether plant cells underwent heat shock. The expression of gfp enabled monitoring of the regeneration process from individual cells to early-stage embryos. Selection of transformed cells was carried out using 150 mg/L kanamycin, while 200 mg/L timentin was added to inhibit bacterial overgrowth. An average of 17 putative transgenic embryos per millilitre of settled cell volume was achieved. Since transgenic plants were not obtained within the project timeframe, molecular characterization could not be conducted. Nevertheless, the results underscore the potential of genetic transformation in taro, an underutilized crop with promising agricultural value.