Canadian Forest Service Publications
Stable Reporter Gene Expression in Western White Pine (Pinus monticola Dougl. ex D. Don) by Agrobacterium-mediated Genetic Transformation. 2010. Liu, J.-J.; Ekramoddoullah, A.K.M. The Americas Journal of Plant Science and Biotechnology. In: Al-Mughrabi, K. (ed) Plant science and biotechnology in North America: Focus on Canada I. Medicinal and Aromatic Plant Science and Biotechnology Volume 4 Special Issue 2: 1-5.
Available from: Pacific Forestry Centre
Catalog ID: 32013
CFS Availability: PDF (request by e-mail)
An Agrobacterium-mediated transformation procedure was developed to transform the mature embryo from Pinus monticola (Dougl. ex D. Don) seeds with two binary vectors containing the reporter gene encoding the green fluorescent protein (GFP) or the ß-glucuronidase protein (GUS), respectively. More than 1000 embryos from independent transformation events were tested for different western white pine seed families. Selection of kanamycin-resistant callus tissues showed that survival rates varied from 33 to 48% in different independent experiments. Transgenic callus tissues survived and continued to grow on the medium with kanamycin (25 µg/mL), whereas non-transgenic callus, regenerated from the embryos of the same seed family, died within 12 weeks. Integration and expression of the introduced reporter gene was confirmed in transgenic western white pine calli by GUS-staining analysis or microscopic observation of GFP fluorescence. Rates for stable reporter gene expression ranged from 2.9 to 6.5% for all embryos co-cultured with Agrobacterium. Our protocol has enabled the routine transformation of western white pine, a species that was previously difficult for gene manipulation. To our knowledge, this is the first report on genetic engineering of this conifer. Our results demonstrate that transgenic gene expression in western white pine is a feasible option for genetic improvement of this valuable conifer as well as for investigating its molecular interactions with the fungal pathogen Cronartium ribicola (J.C. Fisch.).
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