Department of Plant Genetic Transformation, Agricultural Genetic Engineering Research Institute (AGERI), Agriculture Research Center (ARC), Giza, Egypt.
GM Crops Food. 2013 Jan-Mar;4(1):50-7. doi: 10.4161/gmcr.23925. Epub 2013 Jan 1.
Low transformation efficiency and long generation time for production of transgenic Gerbera jemosonii plants leads to vulnerable gene function studies. Thus, transient expression of genes would be an efficient alternative. In this investigation, a transient expression system for gerbera petals based on the Agrobacterium infiltration protocol was developed using the reporter genes β-glucuronidase (gus) and green florescence protein (gfp). Results revealed the incapability of using the gfp gene as a reporter gene for transient expression study in gerbera flowers due to the detection of green fluorescent color in the non-infiltrated gerbera flower petals. However, the gus reporter gene was successfully utilized for optimizing and obtaining the suitable agroinfiltration system in gerbera flowers. The expression of GUS was detectable after three days of agroinfiltration in gerbera cultivars "Express" and "White Grizzly" with dark pink and white flower colors, respectively. The vacuum agroinfiltration protocol has been applied on the cultivar "Express" for evaluating the transient expression of the two genes involved in the anthocyanin pathway (iris-dfr and petunia-f3' 5'h), which is responsible for the color in flowers. In comparison to the control, transient expression results showed change in the anthocyanin pigment in all infiltrated flowers with color genes. Additionally, blue color was detected in the stigma and pollen grains in the infiltrated flowers. Moreover, blue colors with variant intensities were observed in produced calli during the routine work of stable transformation with f3' 5'h gene.
生产转基因非洲菊植株的转化效率低且世代时间长，导致基因功能研究较为脆弱。因此，瞬时表达基因将是一种有效的替代方法。本研究基于农杆菌浸润方案，开发了一种用于非洲菊花瓣的瞬时表达系统，使用报告基因β-葡萄糖醛酸酶（gus）和绿色荧光蛋白（gfp）。结果表明，由于在未浸润的非洲菊花瓣中检测到绿色荧光颜色，因此无法将 GFP 基因用作非洲菊花瞬时表达研究的报告基因。然而，gus 报告基因成功地用于优化和获得非洲菊花中合适的农杆菌浸润系统。在具有深粉色和白色花朵颜色的“Express”和“White Grizzly”两个非洲菊品种中，浸润三天后可检测到 GUS 的表达。真空农杆菌浸润方案已应用于“Express”品种，以评估参与花色苷途径（虹膜-DFR 和矮牵牛-F3'5'H）的两个基因的瞬时表达，花色苷途径决定花的颜色。与对照相比，瞬时表达结果显示，所有带有颜色基因的浸润花朵中的花色苷色素均发生变化。此外，在浸润的花朵中，柱头和花粉粒中检测到蓝色。此外，在使用 F3'5'H 基因进行稳定转化的常规工作中，在产生的愈伤组织中观察到蓝色，其强度不同。
