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Transformation of a recalcitrant grain legume, Vigna mungo L. Hepper, using Agrobacterium tumefaciens-mediated gene transfer to shoot apical meristem cultures

  • Genetic Transformation and Hybridization
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Abstract

The efficiency of Vigna mungo L. Hepper transformation was significantly increased from an average of 1% to 6.5% by using shoot apices excised from embryonic axes precultured on 10 μM benzyl-6-aminopurine (BAP) for 3 days and wounded prior to inoculation in Agrobacterium tumefaciens strain EHA105 carrying the binary vector pCAMBIA2301, which contains a neomycin phosphotransferase gene (nptII) and a β-glucuronidase (GUS) gene (gusA) interrupted by an intron. The transformed green shoots that were selected and rooted on medium containing kanamycin, and which tested positive for nptII gene by polymerase chain reaction, were established in soil to collect seeds. GUS activity was detected in whole T0 shoots and T1 seedlings. All T0 plants were morphologically normal, fertile and the majority of them transmitted transgenes in a 3:1 ratio to their progenies. Southern analysis of T1 plants showed integration of nptII into the plant genome.

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Abbreviations

BAP:

6-Benzylaminopurine

IBA:

Indole-3-butyric acid

2-ip:

6-(γ-γ-Dimethylallylamino) purine

AdS:

Adenine sulfate

MSB:

MS salts with B5 vitamins

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Acknowledgements

P.K. Jaiwal is grateful to the Department of Biotechnology(DBT), New Delhi, for funding the research programme, to The Centre for Application of Molecular Biology to International Agriculture (CAMBIA), Australia for plasmid CAMBIA2301, and to Dr. M.V. Rajam, UDSC, New Delhi, for help in southern hybridisation. Raman Saini is grateful to DBT for a Senior Research Fellowship

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Authors and Affiliations

  1. Department of Biosciences, M.D. University, Rohtak, 124001, India

    Raman Saini & Pawan K. Jaiwal

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

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Communicated by W.A. Parrot

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Saini, R., Jaiwal, P.K. Transformation of a recalcitrant grain legume, Vigna mungo L. Hepper, using Agrobacterium tumefaciens-mediated gene transfer to shoot apical meristem cultures. Plant Cell Rep 24, 164–171 (2005). https://doi.org/10.1007/s00299-005-0934-z

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  • DOI: https://doi.org/10.1007/s00299-005-0934-z

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