Journal of Plant Biochemistry and Biotechnology

, Volume 22, Issue 4, pp 382–391 | Cite as

Factors affecting genetic transformation and shoot organogenesis of Bacopa monnieri (L.) Wettst

  • Diwakar Aggarwal
  • Neha Jaiswal
  • Anil Kumar
  • M. Sudhakara Reddy
Original article


Efficient Agrobacterium tumefaciens mediated T-DNA delivery and subsequent shoot organogenesis has been achieved from Bacopa monnieri. Various factors influenced T-DNA delivery as evident from transient GUS assay. The transient GUS expression was significantly higher (97.7 %) in explants that were pre-cultured before bacterial infection on medium supplemented with 100 μM acetosyringone. Incorporation of acetosyringone into the co-cultivation medium also enhanced transient GUS activity. Explant injury with carborundum paper, co-cultivation period of 2 days and a bacterial density of 0.4 OD600 showed higher transient GUS expression. Following co-cultivation, shoot organogenesis was achieved from leaf segments on basal Murashige and Skoog medium containing 58 mM sucrose. Supplementation of antibiotics (cefotaxime or carbenicillin) at > 250 μg/ml into the medium significantly promoted shoot organogenesis from leaf explants (71.5 % in control and > 83.0 % on medium containing 500 μg/ml of carbenicillin or cefotaxime). Stable transformation of regenerated shoots was confirmed on the basis of GUS activity and PCR amplification of DNA fragments specific to reporter gene (uidA) and selection marker gene (nptII). The expression level of nptII gene in independent transgenic lines was studied using quantitative real time-PCR. Stable transformed shoots after rooting were successfully established in the pots.


Agrobacterium tumefaciens Carbenicillin RT-PCR 16S rRNA 





Neomycin phosphotransferase


Reverse transcriptase-polymerase chain reaction


Real Time - polymerase chain reaction


5-bromo-4-chloro-3-indolyl-β-D-glucuronic acid



Authors are thankful to University Grant Commission (UGC), Govt. of India, New Delhi and Thapar University, Patiala for the financial assistance. Thanks are also due to TIFAC-CORE, Thapar University Patiala for the facilities to carry out this work.


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Copyright information

© Society for Plant Biochemistry and Biotechnology 2012

Authors and Affiliations

  • Diwakar Aggarwal
    • 1
  • Neha Jaiswal
    • 1
  • Anil Kumar
    • 1
  • M. Sudhakara Reddy
    • 1
  1. 1.TIFAC- Centre of Relevance and Excellence in Agro and Industrial Biotechnology, Department of Biotechnology & Environmental SciencesThapar UniversityPatialaIndia

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