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Molecular analysis of transgene and vector backbone integration into the barley genome following Agrobacterium-mediated transformation

  • Genetic Transformation and Hybridization
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Abstract

We report a large-scale study on the frequency of transgene and T-DNA backbone integration following Agrobacterium-mediated transformation of immature barley embryos. One hundred and ninety-one plant lines were regenerated after hygromycin selection and visual selection for GFP expression at the callus stage. Southern blotting performed on a subset of 53 lines that were PCR positive for the GFP gene documented the integration of the GFP gene in 27 of the lines. Twenty-three of these lines expressed GFP in T1 plantlets. Southern blotting with a vector backbone probe revealed that 13 of the 27 lines possessed one or more vector backbone fragments illustrating the regular occurrence of vector backbone integration following Agrobacterium infection of barley immature embryos.

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Abbreviations

GFP:

Green fluorescent protein

nos:

nopaline synthetase

PCR:

Polymerase chain reaction

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Acknowledgements

We thank P. Mathews CSIRO, Canberra for providing us with the vector system and D. Bishop for technical assistance while implementing the procedure. Furthermore the authors would like to thank K. B. Nellerup and O. B. Hansen for their excellent technical support.

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Authors and Affiliations

  1. Department of Genetics and Biotechnology, Danish Institute of Agricultural Sciences, Research Centre Flakkebjerg, Forsoegsvej 1, DK-4200, Slagelse, Denmark

    Mette Lange, Eva Vincze, Marianne G. Møller & Preben B. Holm

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  1. Mette Lange
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  2. Eva Vincze
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  3. Marianne G. Møller
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  4. Preben B. Holm
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Correspondence to Mette Lange.

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Communicated by W. Horwood

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Lange, M., Vincze, E., Møller, M.G. et al. Molecular analysis of transgene and vector backbone integration into the barley genome following Agrobacterium-mediated transformation. Plant Cell Rep 25, 815–820 (2006). https://doi.org/10.1007/s00299-006-0140-7

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  • DOI: https://doi.org/10.1007/s00299-006-0140-7

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