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Over-expression of PIP2;5 aquaporin in a Populus tremula×P.alba clone and its effects on plant responses to low root temperature and osmotic stress

  • Author / Creator
    Ranganathan,Kapilan
  • This dissertation describes the studies that examined: 1. Production of transgenic poplar over-expressing the poplar aquaporin PIP2;5 and 2. Comparison of the physiological properties of a transgenic poplar with the wild-type under different stress conditions including a) short-term low root temperature (LRT), b) prolonged LRT and c) polyethyleneglycol (PEG 6000) induced osmotic stress. Aquaporin PIP2;5 was selected due to its water-transporting capacity and increased expression under water limitation. The coding sequence of PtdPIP2;5 was cloned in the expression vector pCambia1305.2, under the control of maize ubiquitin promoter and the construct was introduced into in vitro grown Populus tremula × P. alba INRA Clone 717-1B4 by Agrobacterium-mediated transformation. Rooted plantlets of transformed poplar were transferred to the growth chamber and transformed lines were confirmed by PCR using primers against the hygromycin resistance gene and quantitative RT-PCR. Two lines exhibiting the highest expression of PIP2;5 were selected and ex vitro plants were generated for experimentation. When the root temperature (RT) was lowered from 20°C to 5°C in solution culture, for 30 minutes, PtdPIP2;5-over-expressing plants had significantly higher net gas exchange rates and root hydraulic conductivities (Lp) compared with the wild-type plants. Recovery was rapid in transgenic lines when the RT was raised back to 20°C. In the transgenic lines, the transcript abundance of PIP2;5 was significantly higher at 20°C and remained highly significant at 5°C for 30 minutes, when compared with the wild-type plants. When the plants were exposed to LRT for 3 weeks in solution culture, transgenic lines showed improved gas exchanging properties and Lp values than the wild-type plants. The transcript abundance of PIP2;5 significantly increased in both transgenic lines at 20°C and PIP2;1 levels increased in one transgenic line, after 3 weeks at 5°C. Transgenic lines had significantly higher growth rates, leaf size, gas exchange parameters and Lp than the wild-type poplars when subjected to PEG 6000 induced osmotic stress, at 50 g L-1. The results suggest that transgenic lines may be more resistant to LRT and osmotic stresses than the wild-type plants. This study confirms the contribution of PIP2;5 on plant water transport, gas exchange and growth.

  • Subjects / Keywords
  • Graduation date
    2015-11
  • Type of Item
    Thesis
  • Degree
    Doctor of Philosophy
  • DOI
    https://doi.org/10.7939/R3251FS1R
  • License
    This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for non-commercial purposes. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.
  • Language
    English
  • Institution
    University of Alberta
  • Degree level
    Doctoral
  • Department
    • Department of Renewable Resources
  • Specialization
    • Forest Biology and Management
  • Supervisor / co-supervisor and their department(s)
    • Janice Cooke (Biological Science)
    • Janusz Zwiazek (Renewable Resources)
  • Examining committee members and their departments
    • Tarriq Siddique (Renewable Resources)
    • Claudio Lovisolo (University of Turin)
    • Barb Thomas (Renewable Resources)