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Characterization of the Populus PtrCesA4 promoter in transgenic Populus alba × P. glandulosa

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Abstract

An efficient shoot regeneration method was developed using leaf explants of Populus alba × P. glandulosa, and the optimized shoot regeneration medium contained 0.1 mg L−1 NAA, 0.5 mg L−1 6-BA and 0.002 mg L−1 TDZ. The factors for Agrobacterium-mediated transformation of this hybrid were further optimized as follows: no preculture step, Agrobacterium cell suspension with an OD600 of 0.6, 20 min infection time and 2d co-cultivation duration. To investigate Populus trichocarpa cellulose synthase A4 promoter (PtrCesA4pro), we constructed the PtrCesA4pro:: β-glucuronidase (GUS) binary vector, in which the cauliflower mosaic virus 35S (CaMV35S) promoter of pBI121was replaced with the PtrCesA4pro to drive the expression of a GUS reporter gene. Forty-eight kanamycin-resistant plantlets were obtained through Agrobacterium-mediated transformation of this hybrid. Genomic DNA PCR and Southern blot analyses confirmed the integration of PtrCesA4pro::GUS into this hybrid and the expression of GUS gene was driven by the PtrCesA4 promoter. A histochemical GUS staining presented the activity of PtrCesA4 promoter in the transgenic lines. No GUS signal was observed in leaves, including the veins and petioles, and notably weak staining was detected in the secondary xylems of the roots. The PtrCesA4 promoter was highly active in the fibers, the vessels of developing xylems, and the fibers of mature phloems. Surprisingly, GUS staining was detected in the cambial cells of the highly lignified stems of these transgenic trees. Our results implicate PtrCesA4 promoter as a good genetic tool for controlling gene function in wood development and tree molecular breeding.

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Acknowledgments

This project was supported by Grants from the National High Technology Research and Development Program of China (2013AA102702), the Program for New Century Excellent Talents in University (NCET-12-0807), the National Key Basic Research Program of China (2012CB114502), and Project of the Natural Sciences Foundation of Heilongjiang Province (Grant C201010).

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Authors and Affiliations

  1. State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, 26 Hexing Road, Harbin, 150040, China

    Guanghui Hai, Zhigang Jia, Wenjing Xu, Chong Wang, Shenquan Cao & Yuxiang Cheng

  2. College of Life Science and Technology, Heilongjiang Bayi Agricultural University, Daqing, 163319, China

    Jinwen Liu

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  1. Guanghui Hai
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  2. Zhigang Jia
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  3. Wenjing Xu
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  4. Chong Wang
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  5. Shenquan Cao
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  6. Jinwen Liu
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  7. Yuxiang Cheng
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Correspondence to Yuxiang Cheng.

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Hai, G., Jia, Z., Xu, W. et al. Characterization of the Populus PtrCesA4 promoter in transgenic Populus alba × P. glandulosa . Plant Cell Tiss Organ Cult 124, 495–505 (2016). https://doi.org/10.1007/s11240-015-0909-x

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  • DOI: https://doi.org/10.1007/s11240-015-0909-x

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