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Plant Cell Reports

, Volume 24, Issue 6, pp 335–340 | Cite as

Development of plant regeneration and transformation protocols for the desiccation-sensitive weeping lovegrass Eragrostis curvula

  • Sandile Ncanana
  • Wolf Brandt
  • George Lindsey
  • Jill Farrant
Cell Biology and Morphogenesis

Abstract

A tissue culture protocol, suitable for transformation, was established for the pasture grass Eragrostis curvula. Callus was generated in the dark from leaf and seed tissues on a medium comprising MS salts supplemented with 2 mg/l 2,4-D, 0.01 mg/l BAP and 2% sucrose. Plant regeneration occurred via organogenesis on the same medium with 6% and 3% sucrose for shoot and root formation, respectively. Optimal regeneration (50 plantlets per callus) occurred when light of 45 μmol/m2 per s was used. The yeast Saccharomyces cerevisiae Hsp12 gene was cloned into the Sac1 of the pCAMBIAUbeeQ vector and callus was transformed by biolistic bombardment. Best transformation (30%) occurred when the target tissue was bombarded twice at a distance of 70 mm using a bombardment pressure of 9,100 kPa. Although successful transformation and transcription of the Hsp12 gene occurred, no Hsp12 protein was found present in tissue extracts of transformed grass.

Keywords

Eragrostis curvula Plant regeneration Transformation Heat shock protein 12 

Abbreviations

BAP

6-Benzylaminopurine

2.4-D

2,4-Dichlorophenoxyacetic acid

GUS

β-Glucuronidase

HSP

Heat shock protein

MS

Murashige Skoog

PCR

Polymerase chain reaction

TDZ

Thidiazuron

X-Gluc

1-Bromo-2-chloro-3-indoyl-β-glucuronic acid

Notes

Acknowledgements

The authors thank Dr S. G. Mundree for donation of the pCAMBIAUbeeQ vector. The work was supported by a grant to Jill Farrant awarded by the National Research Foundation and the DST Innovation fund.

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Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Sandile Ncanana
    • 1
  • Wolf Brandt
    • 1
  • George Lindsey
    • 1
  • Jill Farrant
    • 1
  1. 1.Department of Molecular and Cellular BiologyUniversity of Cape TownPrivate BagSouth Africa

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