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Isopentenyl transferase gene expression offers the positive selection of marker-free transgenic plant of Kalanchoe blossfeldiana

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Abstract

The technologies allowing the production of transgenic plants without selectable marker genes, is of great interest in public and environmental safety. For generating such marker-free transgenic plants, possibility has been offered by Multi-Auto-Transformation [MAT] vector system, which combines positive selection, using the isopentenyl transferase (ipt) gene, with a site-specific recombination that generates marker-free plants. In this study Agrobacterium tumefaciens strain EHA105 harboring an ipt-type MAT vector, pMAT21, containing lacZ, gus genes and the removable cassette in the T-DNA region was used to produce marker-free transgenic Kalanchoe blossfeldiana Poelln., employing ipt gene as the selectable marker gene. Co-cultivated explants were cultured on hormone- and selective agent-free MS medium, and 85% of the regenerated shoots showed ipt-shooty phenotype with GUS expression. Forty-one morphologically normal shoots were produced during the subculture. More than ninety percent of the normal shoots were ipt −, gus − but lacZ + as determined by PCR analyses. These results indicate that the ipt phenotype was clearly distinguishable from non-transgenic as well as transgenic marker-free shoots. This study opens interesting perspective for the generation of marker-free transgenic K. blossfeldiana with objective useful transgene.

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Abbreviations

IPT:

Isopentenyl transferase gene

GUS:

β-Glucuronidase

MS medium:

Murashige and skoog medium

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Acknowledgment

The authors would like to thank Dr. H. Ebinuma, Pulp and Paper Research, Nippon Paper Industries, Tokyo for providing the MAT vector construct.

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Authors and Affiliations

  1. Laboratory of Plant Cell Technology, Graduate School of Horticulture, Chiba University, 648 Matsudo, Matsudo City, Chiba, 271-8510, Japan

    Gunaratnam Thirukkumaran, Raham Sher Khan, Dong Poh Chin, Ikuo Nakamura & Masahiro Mii

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  1. Gunaratnam Thirukkumaran
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  2. Raham Sher Khan
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  3. Dong Poh Chin
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  4. Ikuo Nakamura
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  5. Masahiro Mii
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Correspondence to Masahiro Mii.

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Thirukkumaran, G., Khan, R.S., Chin, D.P. et al. Isopentenyl transferase gene expression offers the positive selection of marker-free transgenic plant of Kalanchoe blossfeldiana . Plant Cell Tiss Organ Cult 97, 237–242 (2009). https://doi.org/10.1007/s11240-009-9519-9

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