Institute of Plant Genetics and Biotechnology, Slovak Academy of Sciences, Akademicka 2, P.O. Box 39A, 95 007, Nitra, Slovak Republic,
Plant Cell Rep. 2015 Mar;34(3):469-81. doi: 10.1007/s00299-014-1726-0. Epub 2014 Dec 13.
Marker-free transgenic plants can be generated with high efficiency by using the Cre/ lox P self-excision system controlled by the pollen- and embryo-specific Arabidopsis DLL promoter. In this work, we aimed to study the feasibility of using the pollen- and embryo-specific DLL promoter of the At4g16160 gene from Arabidopsis thaliana in a Cre/loxP self-excision strategy. A Cre/loxP self-excision cassette controlled by the DLL promoter was introduced into the tobacco genome via Agrobacterium-mediated transformation. No evidence for premature activation of the Cre/loxP system was observed in primary transformants. The efficiency of nptII removal during pollen and embryo development was investigated in transgenic T1 progenies derived from eight self- and four cross-pollinated T0 lines, respectively. Segregation and rooting assays were performed to select recombined T1 plants. Molecular analyses of these plants confirmed the excision event in all analysed T0 lines and marker-free transgenic T1 plants were obtained with efficiency of up to 96.2%. The Arabidopsis DLL promoter appears to be a strong candidate to drive Cre-mediated recombination not only in tobacco as a model plant, but also in other plant species.
利用花粉和胚胎特异性的拟南芥 DLL 启动子控制的 Cre/loxP 自我切除系统，可以高效地产生无标记转基因植物。在这项工作中，我们旨在研究拟南芥 At4g16160 基因的花粉和胚胎特异性 DLL 启动子在 Cre/loxP 自我切除策略中的可行性。通过农杆菌介导的转化，将 Cre/loxP 自我切除盒导入烟草基因组。在初级转化体中未观察到 Cre/loxP 系统的过早激活。在分别来自 8 个自交和 4 个杂交 T0 系的转基因 T1 后代中，研究了在花粉和胚胎发育过程中 nptII 去除的效率。通过分离和生根分析，从 T0 系中选择重组的 T1 植株。对这些植株的分子分析证实了所有分析的 T0 系中都发生了切除事件，并且获得了效率高达 96.2%的无标记转基因 T1 植株。拟南芥 DLL 启动子似乎是一种强有力的候选物，不仅可以在烟草等模式植物中驱动 Cre 介导的重组，也可以在其他植物物种中驱动 Cre 介导的重组。
