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An efficient protocol for regeneration and transformation of Symphyotrichum novi-belgii

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Abstract

A protocol for adventitious shoot formation in Symphyotrichum novi-belgii was developed after investigating the effects of cultivar and hormone combinations. A Murashige and Skoog medium with 1.0 mg l−1 6-benzyladenine induced adventitious shoot formation in 15 out of 19 cultivars. Addition of 0.1 mg l−1 indole-3-acetic acid or naphthaleneacetic acid increased the total number of shoots per explant, but not the number of shoots longer than 1 cm. Addition of dichlorophenoxyacetic acid (2,4-D) promoted callus formation, but inhibited shoot elongation. A transformation system for the two cultivars Victoria Fanny and Victoria Jane was developed by co-cultivation of leaf explants with Agrobacterium tumefaciens. Three bacterial strains (LBA 4404, A281 and C58) all carrying the binary vector, p35S-GUS-INT, and harbouring the uidA gene coding for β-glucuronidase (GUS) were used. Regeneration of transgenic plants after co-cultivation with A281 was independent of cultivar, and all explants produced callus followed by indirect shoot formation. In ‘Victoria Fanny’ shoots were formed faster and without a callus phase after co-cultivation with LBA 4404 or C58. The highest number of potentially transformed shoots was regenerated after co-cultivation of ‘Victoria Fanny’ leaf explants with LBA 4404. Integration of the transgenes in the plant genome was confirmed using PCR and Southern blot hybridisation. To verify that the transgenes could be transferred to offspring, crosses were conducted between three transgenic lines of ‘Victoria Fanny’ and two wild type pollen donors. It was demonstrated that viable seeds were produced and that the uidA gene was inherited.

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Acknowledgments

The authors want to thank Mrs. Elisabeth Kjemtrup-Hansen for expert technical assistance. The research was funded by the Danish Aster Association and The Danish Food Industry Agency under the Ministry of Food, Agriculture and Fisheries (project No. 536-07-654).

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Authors and Affiliations

  1. Department of Food Science, Aarhus University, Kirstinebjergvej 10, 5792, Aarslev, Denmark

    Eline Kirk Mørk, Karin Henriksen, Kell Kristiansen & Karen Koefoed Petersen

  2. Department of Molecular Biology and Genetics, Aarhus University, Forsøgsvej 1, 4200, Slagelse, Denmark

    Henrik Brinch-Pedersen

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  1. Eline Kirk Mørk
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  2. Karin Henriksen
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Correspondence to Eline Kirk Mørk.

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Mørk, E.K., Henriksen, K., Brinch-Pedersen, H. et al. An efficient protocol for regeneration and transformation of Symphyotrichum novi-belgii . Plant Cell Tiss Organ Cult 108, 501–512 (2012). https://doi.org/10.1007/s11240-011-0065-x

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