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Protoplasma

, Volume 251, Issue 3, pp 591–601 | Cite as

An efficient in planta transformation of Jatropha curcas (L.) and multiplication of transformed plants through in vivo grafting

  • Balusamy Jaganath
  • Kondeti Subramanyam
  • Subramanian Mayavan
  • Sivabalan Karthik
  • Dhandapani Elayaraja
  • Rajangam Udayakumar
  • Markandan Manickavasagam
  • Andy GanapathiEmail author
Original Article

Abstract

An efficient and reproducible Agrobacterium-mediated in planta transformation was developed in Jatropha curcas. The various factors affecting J. curcas in planta transformation were optimized, including decapitation, Agrobacterium strain, pin-pricking, vacuum infiltration duration and vacuum pressure. Simple vegetative in vivo cleft grafting method was adopted in the multiplication of transformants without the aid of tissue culture. Among the various parameters evaluated, decapitated plants on pin-pricking and vacuum infiltrated at 250 mmHg for 3 min with the Agrobacterium strain EHA 105 harbouring the binary vector pGA 492 was proved to be efficient in all terms with a transformation efficiency of 62.66 %. Transgene integration was evinced by the GUS histochemical analysis, and the GUS positive plants were subjected to grafting. Putatively transformed J. curcas served as "Scion" and the wild type J. curcas plant severed as "Stock". There was no occurrence of graft rejection and the plants were then confirmed by GUS histochemical analysis, polymerase chain reaction (PCR) and Southern hybridization. Genetic stability of the grafted plants was evaluated by using randomly amplified polymorphic DNA (RAPD), marker which showed 100 % genetic stability between mother and grafted plants. Thus, an efficient in planta transformation and grafting based multiplication of J. curcas was established.

Keywords

Agrobacterium tumefaciens EHA105 Cleft grafting Genetic fidelity In planta transformation PPT 

Abbreviations

bar

Coding region for biolophos resistant gene

CaMV 35S

Cauliflower mosaic virus 35S promoter

gus

β-Glucuronidase

nos

Nopaline synthase terminator

npt II

Neomycin phosotransferase II

RAPD

Randomly amplified polymorphic DNA

UPGMA

Unweighted pair group method with arithmetic mean

Notes

Acknowledgments

The authors are thankful to Bharathidasan University, Tiruchirappalli, Tamil Nadu, India, for providing lab facilities to carry out the present work. The corresponding author is thankful to University Grants Commission (UGC), Govt. of India, for providing fellowship under the UGC–BSR scheme. B. Jaganath is thankful to Bharathidasan University for providing Mookapillai Centenary commemoration endowment fellowship to carry out his Doctoral work. K. Subramanyam is thankful to the Council of Scientific and Industrial Research (CSIR), New Delhi, India for providing Senior Research Fellowship to carry out his Doctoral work.

Conflict of interest

The authors declare that they have no competing interests.

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Copyright information

© Springer-Verlag Wien 2013

Authors and Affiliations

  • Balusamy Jaganath
    • 1
  • Kondeti Subramanyam
    • 1
  • Subramanian Mayavan
    • 1
    • 2
  • Sivabalan Karthik
    • 1
  • Dhandapani Elayaraja
    • 1
  • Rajangam Udayakumar
    • 3
  • Markandan Manickavasagam
    • 1
  • Andy Ganapathi
    • 1
    Email author
  1. 1.Department of Biotechnology and Genetic Engineering, School of BiotechnologyBharathidasan UniversityTamil NaduIndia
  2. 2.Synthetic Biology and Biofuel GroupInternational Center for Genetic Engineering and Biotechnology (ICGEB)New DelhiIndia
  3. 3.Department of BiochemistryGovernment arts college (Autonomous)Tamil NaduIndia

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