Skip to main content

An efficient protocol for particle bombardment-mediated transformation of Centella asiatica callus


In this study, we report an optimization of particle bombardment transformation system for Centella asiatica callus. A total of eight parameters affecting the genetic transformation system were optimized using the synthetic green fluorescent protein (sGFP) as a reporter driven by the CaMV 35S promoter. The results indicated that DNA delivery conditions of 9-cm target distance, 1,100 psi helium pressure, 1.0 μm gold particles size, 27 mmHg chamber vacuum pressure, 2 times number of bombardment, spermidine as precipitation agent, 60 h post-bombardment incubation time and 2 μg plasmid DNA concentration were optimal for C. asiatica callus transformation. The expression of sGFP was monitored using fluorescence microscope and further confirmed using RT-PCR. This optimized genetic transformation system is applicable for rapid transient gene analysis and transgenic C. asiatica production.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4


  1. Brinkhaus B, Linder M, Schuppan D, Hahn EG (2000) Chemical, pharmacological and clinical profile of the East Asian medicinal plant Centella asiatica. Phytomedicine 7:427–448

    PubMed  CAS  Google Scholar 

  2. Kim OT, Bang KH, Shin YS, Lee MJ, Jung SJ, Hyun DY, Kim YC, Seong NS, Cha SW, Hwang B (2007) Enhanced production of asiaticoside from hairy root cultures of Centella asiatica (L.) Urban elicited by methyl jasmonate. Plant Cell Rep 26:1941–1949

    PubMed  Article  CAS  Google Scholar 

  3. Kim OT, Kim HS, Ohyama K, Muranaka T, Choi YE, Lee HY, Kim MY, Hwang B (2010) Upregulation of phytosterol and triterpene biosynthesis in Centella asiatica hairy roots overexpressed ginseng farnesyl diphosphate synthase. Plant Cell Rep 29:403–411

    PubMed  Article  CAS  Google Scholar 

  4. Krishnan VN, Soni KB, Rajmohan K (2008) Agrobacterium tumefaciens mediated genetic transformation in Centella asiatica L. Urban Curr Biotica 2:1–8

    Google Scholar 

  5. Martin KP (2004) Plant regeneration through somatic embryogenesis in medicinally important Centella asiatica L. In Vitro Cell Dev Biol Plant 40:586–591

    Article  CAS  Google Scholar 

  6. Matsuda H, Morikawa T, Ueda H, Yoshikawa M (2001) Medicinal foodstuffs XXVII: saponin constituents Gotu Kola (2) structures of a new ursane- and oleanane-type triterpene oligoglycosides, centellasaponin B, C, and D, from Centella asiatica cultivated in Sri Lanka. Chem Pharm Bull 49:1368–1371

    PubMed  Article  CAS  Google Scholar 

  7. Milen IG, Jost W, Alexandre M (2009) Bioprocessing of plant cell cultures for mass production of targeted compounds. Appl Microbiol Biotechnol 83:809–823

    Article  Google Scholar 

  8. Mook-Jung IH, Shin JE, Yun SH, Huh K, Koh JY, Park HK, Jew SS, Jung MW (1999) Protective effects of asiaticoside derivatives against beta-amyloid neurotoxicity. J Neurosci Res 58:417–425

    PubMed  Article  CAS  Google Scholar 

  9. Murashige T, Skoog F (1962) A revised medium for rapid growth of and bioassay with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  CAS  Google Scholar 

  10. Parveez GKA, Chowdhury MKU, Saleh NM (1997) Physical parameters affecting transient GUS gene expression in oil palm (Elaeis guineensis Jacq.) embryogenic calli via microprojectile bombardment. Ind Crop Prod 8:17–27

    Article  Google Scholar 

  11. Patra A, Rai B, Rout GR, Das P (1998) Successful plant regeneration from callus cultures of Centella asiatica (Linn.) Urban. Plant Growth Regul 24:13–16

    Article  CAS  Google Scholar 

  12. Purkayastha J, Sugla T, Paul A, Solleti SK, Mazumdar P, Basu A, Mohommad A, Ahmed Z, Sahoo L (2010) Efficient in vitro plant regeneration from shoot apices and gene transfer by particle bombardment in Jatropha curcas. Biol Plant 54:13–20

    Article  CAS  Google Scholar 

  13. Rochange F, Serrano L, Marque C, Teulieres C, Boundet AM (1995) DNA delivery into Eucalyptus globules zygotic embryos through biolistics: optimization of the biological and physical parameters of bombardment for two different particle guns. Plant Cell Rep 14:674–678

    Article  Google Scholar 

  14. Southgate EM, Davey MR, Power JB, Marchant R (1995) Factors affecting the genetic engineering of plants by microprojectile bombardment. Biotech Adv 13:631–651

    Article  CAS  Google Scholar 

  15. Sreeramanan S, Maziah M, Abdullah MP, Sariah M, Xavier M, Nor’Aini MF (2005) Physical and biological parameters affecting transient GUS and GFP expression in banana via particle bombardment. Asia Pac J Mol Biol Biotechnol 13:35–57

    Google Scholar 

  16. Suwanaketchanatit C, Piluek J, Peyachoknagul S, Huehne PS (2007) High efficiency of stable genetic transformation in Dendrobium via microprojectile bombardment. Biol Plant 51:720–727

    Article  CAS  Google Scholar 

  17. Tee CS, Maziah M (2005) Optimization of biolistic bombardment parameters for Dendrobium Sonia 17 calluses using GFP and GUS as the reporter system. Plant Cell Tissue Organ Cult 80:77–89

    Article  CAS  Google Scholar 

Download references


Authors will like to thank the members from the laboratory of plant biotechnology and virology for their technical assistance and support.

Author information



Corresponding author

Correspondence to Maziah Mahmood.

Additional information

Communicated by Y. Wang.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Lai, KS., Abdullah, P., Yusoff, K. et al. An efficient protocol for particle bombardment-mediated transformation of Centella asiatica callus. Acta Physiol Plant 33, 2547 (2011).

Download citation


  • Centella asiatica
  • SGFP expression
  • Particle bombardment
  • RT-PCR
  • Transformation