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Stable transformation of mature zygotic embryos and regeneration of transgenic plants of chir pine (Pinus roxbughii Sarg.)

  • Genetic Transformation and Hybridization
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Abstract

A particle inflow gun was used to transfer the plasmid pAHC25 containing the bar gene conferring resistance to glufosinate and the gusA reporter gene, each driven by the maize ubiquitin promoter, to mature embryos of Pinus roxburghii (chir pine). High levels of transient expression were obtained when embryos were cultured for 6 days on 10 μM benzyl adenine-containing medium and then exposed to high osmoticum (0.5 M sucrose) before and after bombardment. Selection on medium containing Basta enabled recovery of stably transformed shoots, both from the epicotyl and from adventitious buds. The primary transformed shoots from the epicotyl were multiplied via axillary shoots. Transformation was confirmed by histochemical staining for β-glucuronidase (GUS) activity, by polymerase chain reaction (PCR) amplification of fragments of gusA and nos terminator, and by the resistance of needles to Basta.

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Abbreviations

GUS :

β-glucuronidase

X-Gluc :

5-bromo-4-choloro-3-indolyl-β-d-glucuronic acid

BA :

benzylaminopurine

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Acknowledgments

The work was supported by Swedish International Development Agency (SIDA), Sweden, and by Department of Biotechnology (DBT), India

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Authors and Affiliations

  1. Tissue Culture Pilot Plant, National Chemical Laboratory, Dr. Homi Bhabha Road, Pashan, Pune, 411 008, Maharashtra, India

    V. A. Parasharami, V. B. Naik & R. S. Nadgauda

  2. Swedish University of Agricultural Sciences, Box 7080, 750 07, Uppsala, Sweden

    S. von Arnold & D. H. Clapham

Authors
  1. V. A. Parasharami
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  2. V. B. Naik
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  3. S. von Arnold
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  4. R. S. Nadgauda
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  5. D. H. Clapham
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Corresponding author

Correspondence to V. A. Parasharami.

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Communicated by M.S. Petersen

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Parasharami, V.A., Naik, V.B., von Arnold, S. et al. Stable transformation of mature zygotic embryos and regeneration of transgenic plants of chir pine (Pinus roxbughii Sarg.). Plant Cell Rep 24, 708–714 (2006). https://doi.org/10.1007/s00299-005-0019-z

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  • DOI: https://doi.org/10.1007/s00299-005-0019-z

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