Efficient Agrobacterium-Mediated Transformation of Hybrid Poplar Populus davidiana Dode x Populus bollena Lauche.
Efficient Agrobacterium-Mediated Transformation of Hybrid Poplar Populus davidiana Dode x Populus bollena Lauche.
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DOI：
10.3390/ijms14022515
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发表时间：
2013-01-25
影响因子：
5.6
通讯作者：
Liu S
中科院分区：
文献类型：
--
作者：
Han X;Ma S;Kong X;Takano T;Liu S
白杨是木本植物离体高效再生的模式生物。我们选择了山杨×大叶杨杂交白杨。通过对Murashige和Skoog培养基中添加0.3 mg/L 6-苄氨基嘌呤和0.08 mg/L萘乙酸的优化，使白杨叶片再生频率达到最高（90%）。提高杨树转化效率对杨树遗传育种和其他应用也具有重要意义。在本研究中，我们发现通过控制叶龄可以显著提高转化频率。通过优化农杆菌浓度（OD 600 = 0.8-1.0）和侵染时间（20-30 min）提高转化效率。透射电镜观察表明，30天龄叶片外植体中的农杆菌入侵数量多于60天和90天龄叶片外植体。利用绿色荧光蛋白（GFP）标记，观察MD-GFP融合蛋白在山杨Dode ×新毛杨叶、茎和根中的表达，以确认转基因整合。Southern和北方杂交分析也显示了T-DNA整合到基因组中和转基因植株的基因表达。我们的研究结果表明，年轻的叶片有更高的转化效率（约30%）比老叶（10%）。
Poplar is a model organism for high in vitro regeneration in woody plants. We have chosen a hybrid poplar Populus davidiana Dode × Populus bollena Lauche. By optimizing the Murashige and Skoog medium with (0.3 mg/L) 6-benzylaminopurine and (0.08 mg/L) naphthaleneacetic acid, we have achieved the highest frequency (90%) for shoot regeneration from poplar leaves. It was also important to improve the transformation efficiency of poplar for genetic breeding and other applications. In this study, we found a significant improvement of the transformation frequency by controlling the leaf age. Transformation efficiency was enhanced by optimizing the Agrobacterium concentration (OD600 = 0.8–1.0) and an infection time (20–30 min). According to transmission electron microscopy observations, there were more Agrobacterium invasions in the 30-day-old leaf explants than in 60-day-old and 90-day-old explants. Using the green fluorescent protein (GFP) marker, the expression of MD–GFP fusion proteins in the leaf, shoot, and root of hybrid poplar P. davidiana Dode × P. bollena Lauche was visualized for confirmation of transgene integration. Southern and Northern blot analysis also showed the integration of T-DNA into the genome and gene expression of transgenic plants. Our results suggest that younger leaves had higher transformation efficiency (~30%) than older leaves (10%).
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影响因子：
3
作者：
Ferreira, Silvia;Batista, Dora;Pais, Maria Salomé
通讯作者：
Pais, Maria Salomé
影响因子：
6.4
作者：
MURASHIGE, T;SKOOG, F
通讯作者：
SKOOG, F
影响因子：
7.2
作者：
Kondou, Youichi;Higuchi, Mieko;Matsui, Minami
通讯作者：
Matsui, Minami
影响因子：
6.2
作者：
Han, KH;Meilan, R;Strauss, SH
通讯作者：
Strauss, SH
影响因子：
3
作者：
Confalonieri, M;Balestrazzi, A;Cella, R
通讯作者：
Cella, R