Biologia plantarum 54:13-20, 2010 | DOI: 10.1007/s10535-010-0003-5

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

J. Purkayastha1, T. Sugla1, A. Paul1, S. K. Solleti2, P. Mazumdar1, A. Basu1, A. Mohommad3, Z. Ahmed3, L. Sahoo1,2,*
1 Center for Energy, Indian Institute of Technology Guwahati, Guwahati, India
2 Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati, India
3 Defence Agricultural Research Laboratory, Haldwani, India

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.

Keywords: benzylaminopurine; β-glucuronidase; gibberellic acid; micropropagation; PCR; Southern blot
Subjects: auxins; cytokinins; ex vitro transfer; gibberellic acid, gibberellin; β-glucuronidase; in vitro culture, direct organogenesis; in vitro culture, rooting; Jatropha curcas; kanamycin; particle bombardment; polymerase chain reaction (PCR); Southern blot analysis; transgenic plants

Received: October 2, 2007; Accepted: July 7, 2008; Published: March 1, 2010Show citation

ACS AIP APA ASA Harvard Chicago IEEE ISO690 MLA NLM Turabian Vancouver
Purkayastha, J., Sugla, T., Paul, A., Solleti, S.K., Mazumdar, P., Basu, A., ... Sahoo, L. (2010). Efficient in vitro plant regeneration from shoot apices and gene transfer by particle bombardment in Jatropha curcas. Biologia plantarum, 54(1), 13-20. doi: 10.1007/s10535-010-0003-5.
Download citation
  • BibTeX (.bib)
  • Bookends (.ris)
  • EasyBib (.ris)
  • EndNote (.enw)
  • EndNote 8 (.xml)
  • ISI WoS (.isi)
  • Medlars (.medlars)
  • Mendeley (.ris)
  • MODS (.xml)
  • Papers (.ris)
  • RefWorks (.txt)
  • RefManager (.ris)
  • RIS (.ris)
  • MS Word (.xml)
  • Zotero (.ris)

    • Open full article

    References

    1. Altpeter, F., Vasil, V., Srivastava, V., Stoger, E., Vasil, I.K.: Accelerated production of transgenic wheat (Triticum aestivum L.) plants. - Plant Cell Rep. 16: 12-17, 1996. Go to original source...
    2. Aragao, F.J.L., Rech E.L.: Morphological factors influencing recovery of transgenic bean plants (Phaseolus vulgaris L.) of a carioca cultivar. - Int. J. Plant Sci. 158: 157-163, 1997.
    3. Baskaran, P., Jayabalan, N.: An efficient micropropagation system for Eclipta alba - avaluable medicinal herb. - In Vitro cell. dev. Biol. Plant. 41: 532-539, 2005. Go to original source...
    4. Brassard, N., Brissette, L., Lord, D., Laliberte, S.: Elongation, rooting and acclimatization of micropropagated shoots from mature material of hybrid larch. - Plant Cell Tissue Organ Cult. 44: 37-44, 1996. Go to original source...
    5. Bregitzer, P., Tonks, D.: Inheritance and expression of transgenes in barley. - Crop Sci. 43: 4-12, 2003. Go to original source...
    6. Choi, H.W., Lemaux, P.G., Cho, M.J.: Increased chromosomal variation in transgenic versus nontransgenic barley (Hordeum vulgare L.) plants. - Crop Sci. 40: 524-533, 2000. Go to original source...
    7. Christou, P.: Genetic transformation of crop plants using microprojectile bombardment. - Plant J. 2: 275-281, 1992. Go to original source...
    8. Datla, R.S.S., Hammerlindl, J.K., Pelcher, L.E., Crosby, W.L., Selvaraj, G.: A bifunctional fusion between β-glucuronidase and neomycin phosphotransferase: a broad-spectrum marker enzyme for plants. - Gene 101: 239-246, 1991. Go to original source...
    9. Figueiredo, S.F.L., Albarello, N., Viana, V.R.C.: Micropropagation of Rollinia mucosa (Jacq.) Baill. - In Vitro cell. Dev. Biol. Plant. 37: 471-475, 2001. Go to original source...
    10. Folling, L., Olesen, A.: Transformation of wheat (Triticum aestivum L.) microspore-derived callus and microspores by particle bombardment. - Plant Cell Rep. 20: 1098-1105, 2002. Go to original source...
    11. Fu, X., Duc, L.T., Fontana, S., Bong, B.B., Tinjuangjun, P., Sudhakar, D., Twyman, R.M., Christou, P., Kohli, A.: Linear transgene constructs lacking vector backbone sequences generate low-copy-number transgenic plants with simple integration patterns. - Transgenic Res. 9: 11-19, 2000. Go to original source...
    12. Geetha, N., Venkatachalam, P., Prakash, V., Sita Lakshmi, G.: High frequency induction of multiple shoots and plant regeneration from seedling explants of pigeonpea (Cajanus cajan). - Curr Sci. 75: 1036-1040, 1998.
    13. Gopi, C., Vatsala, T.M., Ponmurugan, P.: In vitro multiple shoot proliferation and plant regeneration of Vanilla planifolia Andr. - a commercial spicy orchid. - J. Plant Biotechnol. 8: 37-41, 2006.
    14. Heller, J.: Physic Nut, Jatropha curcas L. Promoting the Conservation and Use of Underutilized and Neglected Crops. - International Plant Genetic Resource Institute, Rome 1996.
    15. Hossain, M., Islam, R., Karim, M.R., Joarder, O.I., Biswas, B.K.: Regeneration of plantlets from in vitro cultured cotyledons of Aegle marmelos Corr. (Rutaceae). - Sci. Hort. 57: 315-321, 1994. Go to original source...
    16. Iglesias, V.A., Gisel, A., Bilang, R., Leduc, N., Potrykus, I., Sautter, C.: Transient expression of visible marker genes in meristem cells of wheat embryos after ballistic microtargetting. - Planta 192: 84-91, 1994.
    17. Jayaraj, J., Liang, G.H., Muthukrishnan, S., Punja, Z.K.: Generation of low copy number and stably expressing transgenic bentgrass plants using minimal gene casette bombardment. - Biol. Plant. 52: 215-221, 2008. Go to original source...
    18. Jefferson, R.A., Kavanagh, T.A., Bevan, M.W.: GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. - EMBO J. 6: 3901-3907, 1987. Go to original source...
    19. Jha, T.B., Mukherjee, P., Datta, M.M.: Somatic embryogenesis in Jatropha curcas Linn., an important biofuel plant. - Plant Biotechnol. Rep. 1: 135-140, 2007. Go to original source...
    20. Johri, B.M., Srivastava, P.S.: Morphogenesis in endoserm cultures. - Z. Pflanzenphysiol. 70: 285-304, 1973. Go to original source...
    21. Jordan, M., Oyanedel, E.: Regeneration of Pouteria lucuma (Sapotaceae) plants in vitro. - Plant Cell Tissue Organ Cult. 31: 249-252, 1992.
    22. Kamo, K., Blowers, A.: Tissue specificity and expression level of gusA under rolD, mannopine synthase and translation elongation factor 1 subunit promoters in transgenic Gladiolus plants. - Plant Cell Rep. 18: 809-815, 1999. Go to original source...
    23. Kaushik, N., Kumar, K., Kumar, S., Kaushik, N., Roy, S.: Genetic variability and divergence studies in seed traits and oil content of jatropha (Jatropha curcas L.) accessions. - Biomass Bioener. 31: 497-502, 2007. Go to original source...
    24. Lee, C.W., Yeckes, J., Thomas, J.C.: Tissue culture propagation of Euphorbia lathyris and Asclepias erosa. - Hort. Sci. 17: 533, 1982.
    25. Murashige, T., Skoog, F.: A revised medium for rapid growth and bioassays with tobacco tissue cultures. - Physiol. Plant. 15: 473-497, 1962. Go to original source...
    26. Naik, S.K., Pattnaik, S., Chand, P.K.: High frequency axillary shoot proliferation and plant regeneration from cotyledonary nodes of pomegranate (Punica granatum L.). - Sci. Hort. 85: 261-270, 2000. Go to original source...
    27. Nair, N.G., Kartha, K.K., Gamborg, O.L.: Effect of growth regulators on plant regeneration from shoot apical meristems of cassava (Manihot esculenta Crantz) and on the culture of internodes in vitro. - Z. Pflanzenphysiol. 95: 51-56, 1979. Go to original source...
    28. Ortiz, J.P.A., Reggiardo, M.I., Ravizzini, R.A., Altabe, S.G., Cervigni, G.D.L., Spitteler, M.A., Morata, M.M., Elias, F.E, Vallejos, R.H.: Hygromycin resistance as an efficient selectable marker for wheat stable transformation. - Plant Cell Rep. 15: 877-881, 1996. Go to original source...
    29. Parveez, G.K.A., Chowdhury, M.K.U., Saleh, N.M. Physical parameters affecting transient GUS gene expression in oil palm (Elaeis guineensis Jacq.) using the biolistic device. - Industr. Crops Prod. 6: 41-50, 1997. Go to original source...
    30. Perl, A., Kless, H., Blumenthal, A., Galili, G., Galunj, E.: Improvement of plant regeneration and GUS expression in scutellar wheat calli by optimization. - Mol. gen. Genet. 235: 279-284, 1992. Go to original source...
    31. Purohit, S.D., Singhvi, A.: Micropropagation of Achras sapota through enhanced axillary branching. - Sci. Hort. 76: 219-229, 1998. Go to original source...
    32. Ramesh, M., Gupta, A.K.: Transient expression of β-glucuronidase gene in indica and japonica rice (Oryza sativa L.) callus cultures after different stages of co-bombardment. - Afr. J. Biotechnol. 4: 596-600, 2005.
    33. Ripley, K.P., Preece, J.E.: Micropropagation of Euphorbia lathyrus L. - Plant Cell Tissue Org. Cult. 5: 213-218, 1986. Go to original source...
    34. Rogers, S.O., Benedich, A.J.: Extraction of DNA from plant tissues. In: Gelvin, S.B., Schilperoot, R.A. (ed.): Plant Molecular Biology Manual. Pp. A6/1-10. Kluwer Academic Publishers, Dordrecht 1988.
    35. Schopke, C., Taylor, N.J., Carcamo, R., Beachy, R.N.: Optimization of parameters for particle bombardment of embryogenic suspension cultures of cassava (Manihot esculenta Crantz) using computer image analysis. - Plant Cell Rep. 16: 526-530, 1997. Go to original source...
    36. Shekhawat, N.S., Rathore, T.S., Singh, N.S., Deora, S.R.R.: Factors affecting In vitro clonal propagation of Prosopsis cineraria. - Plant Growth Regul. 12: 273-280, 1993. Go to original source...
    37. Srivastava, P.S.: In vitro induction of triploid roots and shoots from mature endosperm of Jatropha panduraefolia. - Z. Pflanzenphysiol. 66: 93-96, 1971. Go to original source...
    38. Srivastava, P.S., Johri, B.M.: Morphogensis in mature endosperm cultures of Jatropha panduraefolia Beitr. - Biol. Pflanz. 50: 255-268, 1974.
    39. Sticklen, M.B., Oraby, H.F.: Shoot apical meristem: a sustainable explant for genetic transformation of cereal crops. - In Vitro cell. dev. Biol. Plant. 41: 187-200, 2005. Go to original source...
    40. Sujatha, M., Dhingra, M.: Rapid plant regeneration from various explants of Jatropha integerrima. - Plant Cell Tissue Organ Cult. 35: 293-296, 1993. Go to original source...
    41. Sujatha, M., Makkar, H.P.S., Becker, K.: Shoot bud proliferation from axillary nodes and leaf sections of nontoxic Jatropha curcas L. - Plant Growth Regul. 47: 83-90, 2005. Go to original source...
    42. Sujatha, M., Mukta, N.: Morphogenesis and plant regeneration from tissue cultures of Jatropha curcas. - Plant Cell Tissue Organ Cult. 44: 135-141, 1996. Go to original source...
    43. Sujatha, M., Reddy, T.P.: Differential cytokinin effects on the stimulation of in vitro shoot proliferation from meristematic explants of castor (Ricinus communis L.). - Plant Cell Rep. 17: 561-566, 1998. Go to original source...
    44. Suwanaketchanatit, C., Piluek, J., Peyachoknagul, S., Huehne, P.S.: High efficiency of stable genetic transformation in Dendrobium via microprojectile bombardment. - Biol. Plant. 51: 720-727, 2007. Go to original source...
    45. Tideman, J., Hawker, J.S.: In vitro propagation of latex producing plants. - Ann. Bot. 49: 273-279, 1982. Go to original source...
    46. Tiwari, A.K., Kumar, A., Raheman, H.: Biodiesel production from jatropha oil (Jatropha curcas) with high free fatty acids: an optimized process. - Biomass Bioener. 31: 569-575, 2007. Go to original source...
    47. Vengadesan, G., Ganapathi, A., Prem Anand, R., Anbazhagan, V.R.: In vitro propagation of Acacia sinuata (Lour.) Merr. via cotyledonary nodes. - Agroforestry Syst. 55: 9-15, 2002. Go to original source...
    48. Xiao, L., Ha, S.B.: Efficient selection and regeneration of creeping bentgrass transformants following particle bombardment. - Plant Cell Rep. 16: 874-878, 1997. Go to original source...
    49. Zhang, W., Chen, L., Qu, R., Marmey, P., Beachy, R., Fauquet, C.: Regeneration of fertile transgenic indica (group 1) rice plants following microprojectile transformation of embryogenic suspension culture cells. - Plant Cell Rep. 15: 465-469, 1996. Go to original source...

    Return to the content