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Optimization in Agrobacterium-mediated transformation of Anthurium andraeanum using GFP as a reporter Qing Zhao Ji Jing* Gang Wang* Jie Hua Wang Yuan Yuan Feng Han Wen Xing Chun Feng Guan *Corresponding author Financial support: The research was supported by Transgenic Anthurium Flowers, Butterfly Orchid Breeding Techniques and Rapid Propagation Technology of the Tianjin Science and Technology Commission (Grant no. 06YFGZNC01700) and National GMO Major Projects (Grant no. 2009ZX08003-019B). Keywords: cocultivation, gene transfer, green fluorescent protein, regeneration.
Although Agrobacterium-mediated transformation protocols for many economically important plant species have been well established, protocol for a number of flowering plants including Anthurium andraeanum remains challenging. In this study, we report success in generating transgenic Anthurium andraeanum cv Arizona using Agrobacterium GV3101 strain harboring a binary vector carrying gfp as a reporter gene. The possibility of facilitating the screening process for transgenic plants expressing functional proteins using gfp marker was explored. In order to realize high transformation efficiency, different explant sources including undifferentiated callus pieces and petioles were compared for their regeneration efficiency and susceptibility to Agrobacterium-mediated transformation. We also optimized the concentration of AS added to co-cultivation media. Genomic PCR revealed that 11 of the 22 resistant plantlets regenerated on selective medium were successfully transformed. Green fluorescence was observed using a fluorescence microscope in 7 of the 11 PCR-positive plants, indicating GFP was expressed stably in the transformed Anthurium andraeanum. The highest transformation efficiency obtained in this study was 1.71% (percentage of explants with transgenic shoots in total explants) when callus explants were used as starting material and 125 μmol l-1 AS was added during the co-cultivation process. |