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Agrobacterium tumefaciens mediated transfer of Phaseolus vulgaris α-amylase inhibitor-1 gene into mungbean Vigna radiata (L.) Wilczek using bar as selectable marker

  • Genetic Transformation and Hybridization
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Abstract

Morphologically normal and fertile transgenic plants of mungbean with two transgenes, bar and α-amylase inhibitor, have been developed for the first time. Cotyledonary node explants were transformed by cocultivation with Agrobacterium tumefaciens strain EHA105 harboring a binary vector pKSB that carried bialaphos resistance (bar) gene and Phaseolus vulgaris α-amylase inhibitor-1 (αAI-1) gene. Green transformed shoots were regenerated and rooted on medium containing phosphinothricin (PPT). Preculture and wounding of the explants, presence of acetosyringone and PPT-based selection of transformants played significant role in enhancing transformation frequency. Presence and expression of the bar gene in primary transformants was evidenced by PCR-Southern analysis and PPT leaf paint assay, respectively. Integration of the Phaseolus vulgaris α-amylase inhibitor gene was confirmed by Southern blot analysis. PCR analysis revealed inheritance of both the transgenes in most of the T1 lines. Tolerance to herbicide was evidenced from seed germination test and chlorophenol red assay in T1 plants. Transgenic plants could be recovered after 8–10 weeks of cocultivation with Agrobacterium. An overall transformation frequency of 1.51% was achieved.

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Abbreviations

BAP:

6-Benzylaminopurine

IBA:

Indole-3-butyric acid

B5 :

Gamborg's medium (1968)

bar :

Bialaphos resistance

αAI :

α-Amylase inhibitor

PPT:

Phosphinothricin.

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Acknowledgements

We are grateful to the Department of Science and Technology, New Delhi for financial assistance. We are also thankful to Prof. M. J. Chrispeels, University of California, USA for the plasmid KSB harboring α-amylase inhibitor-1 and bar genes and to Prof. Anil Grover, University of Delhi South Campus, New Delhi for providing laboratory facilities for molecular biology work.

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Authors and Affiliations

  1. Advanced Centre for Biotechnology, M. D. University, Rohtak, 124001, India

    Sonia, Raman Saini & Pawan K. Jaiwal

  2. Department of Environmental Sciences, B. B. Ambedkar University, Raebareli Road, Lucknow, 226025, India

    Rana P. Singh

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  1. Sonia
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  2. Raman Saini
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  4. Pawan K. Jaiwal
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Correspondence to Pawan K. Jaiwal.

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Communicated by D. A. Somers

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Sonia, Saini, R., Singh, R.P. et al. Agrobacterium tumefaciens mediated transfer of Phaseolus vulgaris α-amylase inhibitor-1 gene into mungbean Vigna radiata (L.) Wilczek using bar as selectable marker. Plant Cell Rep 26, 187–198 (2007). https://doi.org/10.1007/s00299-006-0224-4

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  • DOI: https://doi.org/10.1007/s00299-006-0224-4

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