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Biologia Plantarum

, Volume 54, Issue 1, pp 13–20 | Cite as

Efficient in vitro plant regeneration from shoot apices and gene transfer by particle bombardment in Jatropha curcas

  • J. Purkayastha
  • T. Sugla
  • A. Paul
  • S. K. Solleti
  • P. Mazumdar
  • A. Basu
  • A. Mohommad
  • Z. Ahmed
  • L. SahooEmail author
Original Papers

Abstract

An efficient and reproducible in vitro plant regeneration system from shoot apices was developed in Jatropha curcas. Benzylaminopurine (BAP; 2.5 μM) was most effective in inducing an average of 6.2 shoots per shoot apex. Incorporation of gibberellic acid (GA3; 0.5 μM) to basal medium was found essential for elongation of shoots. The BAP-habituated mother explants continuously produced shoots during successive subculture without any loss of morphogenic potential. The shoots rooted efficiently on half-strength MS medium. The rooted plantlets were acclimatized with more than 98 % success and the plants transferred to soil:compost in nursery showed no sign of variation compared to the seed-grown plants. The whole process of culture initiation to plant establishment was accomplished within 5–6 weeks. A genetic transformation system in J. curcas was established for the first time, using bombardment of particles coated with plasmid pBI426 with a GUS-NPT II fusion protein under the control of a double 35S cauliflower mosaic virus (CaMV) promoter. The β-glucuronidase (GUS) activity in J. curcas shoot apices was significantly affected by the gold particle size, bombardment pressure, target distance, macrocarrier travel distance, number of bombardments, and type and duration of osmotic pre-treatment. The proliferating bombarded shoot apices were screened on medium supplemented with 25 mg dm−3 kanamycin and surviving shoots were rooted on medium devoid of kanamycin. The integration of the transgene into genomic DNA of transgenic plants was confirmed by PCR and Southern blot hybridization. The transgenic plants showed insertion of single to multiple copies of the transgene.

Additional key words

benzylaminopurine β-glucuronidase gibberellic acid micropropagation PCR Southern blot 

Abbreviations

BAP

6-benzylaminopurine

GA3

gibberellic acid

GUS

β-glucuronidase

NAA

α- naphthaleneacetic acid

IBA

indole-3-butyric acid

MS

Murashige and Skoog’s (1962) medium

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Notes

Acknowledgements

The authors thank NRC (PBI), Canada for pBI426 plasmid, and NEDFi Guwahati and DARL Pithoragarh India for financial assistance.

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Copyright information

© Springer Science+Business Media B.V. 2010

Authors and Affiliations

  • J. Purkayastha
    • 1
  • T. Sugla
    • 1
  • A. Paul
    • 1
  • S. K. Solleti
    • 2
  • P. Mazumdar
    • 1
  • A. Basu
    • 1
  • A. Mohommad
    • 3
  • Z. Ahmed
    • 3
  • L. Sahoo
    • 1
    • 2
    Email author
  1. 1.Center for EnergyIndian Institute of Technology GuwahatiGuwahatiIndia
  2. 2.Department of BiotechnologyIndian Institute of Technology GuwahatiGuwahatiIndia
  3. 3.Defence Agricultural Research LaboratoryHaldwaniIndia

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