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Efficient and stable transgene suppression via RNAi in field-grown poplars

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Abstract

The efficiency and stability of RNA interference (RNAi) in perennial species, particularly in natural environments, is poorly understood. We studied 56 independent poplar RNAi transgenic events in the field over 2 years. A resident BAR transgene was targeted with two different types of RNAi constructs: a 475-bp IR of the promoter sequence and a 275-bp IR of the coding sequence, each with and without the presence of flanking matrix attachment regions (MARs). RNAi directed at the coding sequence was a strong inducer of gene silencing; 80% of the transgenic events showed more than 90% suppression. In contrast, RNAi targeting the promoter resulted in only 6% of transgenic events showing more than 90% suppression. The degree of suppression varied widely but was highly stable in each event over 2 years in the field, and had no association with insert copy number or the presence of MARs. RNAi remained stable during a winter to summer seasonal cycle, a time when expression of the targeted transgene driven by an rbcS promoter varied widely. When strong gene suppression was induced by an IR directed at the promoter sequence, it was accompanied by methylation of the homologous promoter region. DNA methylation was also observed in the coding region of highly suppressed events containing an IR directed at the coding sequence; however, the methylation degree and pattern varied widely among those suppressed events. Our results suggest that RNAi can be highly effective for functional genomics and biotechnology of perennial plants.

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Acknowledgements

We would like to thank Plant Genetic Systems for kindly providing the vector pTTM8, Monsanto for the use of their CP4 gene, Dr. Steven Spiker for providing MAR fragment, and Dr. Peter Waterhouse for the use of pHANNIBAL. This work was supported by Initiative for Future Agriculture and Food Systems (award # 00-52100-9623) from the USDA Cooperative State Research, Education, and Extension Service; TBGRC/TGERC Research Cooperative; the OSU Foundation Forest Biotechnology Fund; and the Conrad Wessela Scholarship Fund at the OSU College of Forestry.

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  1. Jingyi Li

    Present address: Department of Plant and Microbial Biology, The University of California, Berkeley, CA, 94720-3102, USA

Authors and Affiliations

  1. Department of Forest Science, Oregon State University, Corvallis, OR, 97331-5752, USA

    Jingyi Li, Olga Shevchenko, Cathleen Ma & Steven H. Strauss

  2. Department of Forestry, Virginia Polytechnic Institute and State University, Blacksburg, VA, 24061-0324, USA

    Amy M. Brunner

  3. Department of Forestry and Natural Resources, Purdue University, West Lafayette, IN, 47907-2061, USA

    Richard Meilan

  4. Molecular Assisted Breeding Laboratory, Nunhems USA, Inc. , 8850 59th Ave. NE, Brooks, OR, 97305, USA

    Jeffrey S. Skinner

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  1. Jingyi Li
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Correspondence to Steven H. Strauss.

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Li, J., Brunner, A.M., Shevchenko, O. et al. Efficient and stable transgene suppression via RNAi in field-grown poplars. Transgenic Res 17, 679–694 (2008). https://doi.org/10.1007/s11248-007-9148-1

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