Gene transfer into Solanum tuberosum via Rhizobium spp.
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ORIGINAL PAPER
Gene transfer into Solanum tuberosum via Rhizobium spp. Toni Wendt • Fiona Doohan Dominik Winckelmann • Ewen Mullins
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Received: 21 December 2009 / Accepted: 15 June 2010 / Published online: 27 June 2010 Ó Springer Science+Business Media B.V. 2010
Abstract Agrobacterium tumefaciens-mediated transformation (ATMT) is the preferred technique for gene transfer into crops. A major disadvantage of the technology remains the complexity of the patent landscape that surrounds ATMT which restricts its use for commercial applications. An alternative system has been described (Broothaerts et al. in Nature 433:629-633, 2005) detailing the propensity of three rhizobia to transform the model crop Arabidopsis thaliana, the non-food crop Nicotiana tabacum and, at a very low frequency, the monocotyledonous crop Oryza sativa. In this report we describe for the first time the genetic transformation of Solanum tuberosum using the non-Agrobacterium species Sinorhizobium meliloti, Rhizobium sp. NGR234 and Mesorhizobium loti. This was achieved by combining an optimal bacterium and host co-cultivation period with a low antibiotic regime during the callus and shoot induction stages. Using this optimized protocol the
Electronic supplementary material The online version of this article (doi:10.1007/s11248-010-9423-4) contains supplementary material, which is available to authorized users. T. Wendt D. Winckelmann E. Mullins (&) Teagasc Crops Research Centre, Plant Biotechnology Unit, Oak Park, Carlow, Ireland e-mail: [email protected] T. Wendt F. Doohan School of Biology and Environmental Sciences, University College Dublin, Belfield, Dublin 4, Ireland
transformation frequency (calculated as % of shoots equipped with root systems with the ability to grow in rooting media supplemented with 25 lg/ml hygromycin) of the rhizobia strains was calculated at 4.72, 5.85 and 1.86% for S. meliloti, R. sp. NGR234 and M. loti respectively, compared to 47.6% for the A. tumefaciens control. Stable transgene integration and expression was confirmed via southern hybridisation, quantitative PCR analysis and histochemical screening of both leaf and/or tuber tissue. In light of the rapid advances in potato genomics, combined with the sequencing of the potato genome, the ability of alternative bacteria species to genetically transform this major food crop will provide a novel resource to the Solanaceae community as it continues to develop potato as both a food and non-food crop. Keywords Non-Agrobacterium strains Genetic transformation Solanum tuberosum Potato Transbacter Rhizobia
Introduction Potato (Solanum tuberosum L.) is the world’s third most important food crop with over 325 million tonnes produced in 2007 (FAOSTAT, 2009). As the global population advances to 9 billion by 2050, the potato will continue to play a significant role in securing global food reserves (U.N. Secretariat 2007). In parallel to its importance as a stable food source,
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the traditional remit of the potato has been enhanced and expande
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