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Int. J. Mol. Sci. 2013, 14(2), 2515-2528; doi:10.3390/ijms14022515
Article

Efficient Agrobacterium-Mediated Transformation of Hybrid Poplar Populus davidiana Dode × Populus bollena Lauche

1
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1
,
1,2
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3
 and
1,*
1 Key Laboratory of Saline-alkali Vegetation Ecology Restoration in Oil Field (SAVER), Ministry of Education, Alkali Soil Natural Environmental Science Center (ASNESC), Northeast Forestry University, Harbin Hexing Road, Harbin 150040, China 2 Institute of Microbiology, Heilongjiang Academy of Sciences, Harbin 150010, China 3 Asian Natural Environmental Science Center, University of Tokyo, Nishitokyo-shi, Tokyo 188-0002, Japan
* Author to whom correspondence should be addressed.
Received: 7 November 2012 / Revised: 29 December 2012 / Accepted: 17 January 2013 / Published: 25 January 2013
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Abstract

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%).
Keywords: regeneration; transformation; leaf age; poplar; Agrobacterium regeneration; transformation; leaf age; poplar; Agrobacterium
This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).
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Han, X.; Ma, S.; Kong, X.; Takano, T.; Liu, S. Efficient Agrobacterium-Mediated Transformation of Hybrid Poplar Populus davidiana Dode × Populus bollena Lauche. Int. J. Mol. Sci. 2013, 14, 2515-2528.

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