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A passage through in vitro culture leads to efficient production of marker-free transgenic plants in Brassica juncea using the Cre–loxP system

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Abstract

The Cre–loxP site-specific recombination system was deployed for removal of marker genes from Brassica juncea (Indian mustard). Excision frequencies, monitored by removal of nptII or gfp genes in F1 plants of crosses between LOX and CRE lines, were high in quiescent, differentiated somatic tissues but extremely poor in the meristematic regions (and consequently the germinal cells) thus preventing identification and selection of marker-free transgenic events which are devoid of both the marker gene as well as the cre gene, in F2 progeny. We show that a passage through in vitro culture of F1 leaf explants allows efficient development of marker-free transgenics in the F2 generation addressing current limitations associated with efficient use of the Cre/loxP technology for marker gene removal.

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Acknowledgements

Financial support for this research was provided by DOFCO, a subsidiary of the National Dairy Development Board (NDDB). B. S. Yadav provided technical support. The gfp gene was a kind gift from Prof Jen Sheen. We are grateful to the Department of Botany, University of Delhi for extending use of the confocal microscope facility.

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  1. N. Arumugam

    Present address: Department of Biotechnology, Pondicherry University, Pondicherry, 605 014, India

Authors and Affiliations

  1. Centre for Genetic Manipulation of Crop Plants, University of Delhi South Campus, Benito Juarez Road, New Delhi, 110 021, India

    N. Arumugam, Vibha Gupta, Arun Jagannath, Arundhati Mukhopadhyay, Akshay K. Pradhan & Deepak Pental

  2. Department of Biotechnology, Pondicherry University, Pondicherry, 605 014, India

    Akshay K. Pradhan, Pradeep Kumar Burma & Deepak Pental

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  1. N. Arumugam
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  2. Vibha Gupta
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  3. Arun Jagannath
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  4. Arundhati Mukhopadhyay
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  7. Deepak Pental
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Correspondence to Deepak Pental.

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N. Arumugam and Vibha Gupta have contributed equally to this work.

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Arumugam, N., Gupta, V., Jagannath, A. et al. A passage through in vitro culture leads to efficient production of marker-free transgenic plants in Brassica juncea using the Cre–loxP system. Transgenic Res 16, 703–712 (2007). https://doi.org/10.1007/s11248-006-9058-7

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