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Protamine-mediated DNA coating remarkably improves bombardment transformation efficiency in plant cells

Plant Cell Reports volume 28, pages 213–221 (2009)Cite this article

Abstract

We have developed a method by which remarkably higher efficiencies of transient and stable transformation were achieved in bombardment transformation of plants. Over fivefold increase in transient gus gene expression was achieved when rice or maize suspension cells were bombarded with gold particles coated with plasmid DNA in the presence of protamine instead of the conventional spermidine. A 3.3-fold improvement in stable transformation efficiency was also observed using rice suspension cells with the new coating approach. The coated protamine-plasmid DNA complex resisted degradation by a DNase or by rice cell extract much longer than the spermidine-plasmid DNA complex. The results from this study suggest that protamine protects plasmid DNA longer than spermidine when being delivered inside the cells, probably by forming a nano-scale complex, and thus helps improve the efficiency of particle bombardment-mediated plant transformation.

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Abbreviations

GUS:

β-Glucuronidase

hyg B:

Hygromycin B

LUC:

Luciferase

MU:

Methylumbelliferone

MUG:

4-Methylumbelliferyl-β-d-glucuronide

NOS:

Nopaline synthase

pDNA:

Plasmid DNA

X-Gluc:

5-Bromo-4-chloro-3-indolyl β-d-glucuronic acid

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Acknowledgments

The Plasmid DNA construct pJD313 from Dr. J. Sheen, and technical assistance from S. Srilaxmi are gratefully acknowledged. The authors are also thankful to Dr. A. Weissinger for helpful discussion.

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Author notes

  1. Elumalai Sivamani

    Present address: Syngenta Biotechnology, Inc., Research Triangle Park, NC, 27709, USA

Authors and Affiliations

  1. Department of Crop Science, North Carolina State University, Raleigh, NC, 27695-7620, USA

    Elumalai Sivamani & Rongda Qu

  2. Departments of Chemistry and Biomedical Sciences, Missouri State University, 901 South National Avenue, Springfield, MO, 65897, USA

    Robert K. DeLong

Authors
  1. Elumalai Sivamani
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  2. Robert K. DeLong
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  3. Rongda Qu
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Corresponding author

Correspondence to Rongda Qu.

Additional information

Communicated by P. Ozias Akins.

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Sivamani, E., DeLong, R.K. & Qu, R. Protamine-mediated DNA coating remarkably improves bombardment transformation efficiency in plant cells. Plant Cell Rep 28, 213–221 (2009). https://doi.org/10.1007/s00299-008-0636-4

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  • DOI: https://doi.org/10.1007/s00299-008-0636-4

Keywords

  • DNA coating
  • DNA delivery
  • GUS
  • Plant transformation
  • Particle bombardment
  • Protamine