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Gene delivery using microinjection of agrobacterium to embryonic shoot apical meristem of elite indica rice cultivars

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Abstract

A method of gene transfer to elite indica rice cultivars (BPT5204, IW-Ponni and CR1009) by microinjecting the embryonic shoot apical meristem (ESAM) of seeds with Agrobacterium is described. The ESAM microinjected with Agrobacterium cells containing binary plasmid was incubated in co-cultivation media at 28°C for 3 days in the dark and subsequently transferred to selection medium with appropriate supplements and antibiotic. The frequency of gene transfer under various concentrations of Agrobacterium and acetosyringone used were analyzed by staining for the presence of the marker enzyme β-glucuronidase (GUS) carried as part of the binary plasmid and growth in the presence of antibiotic hygromycin. Results of this investigation show that the maximum efficiency of gene transfer took place when Agrobacterium was used at an OD600 of 0.3 and grown in the presence of 20 mg 1−1 acetosyringone.

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Fig. 1
Fig. 2

Abbreviations

BAP:

benzylaminopurine

ESAM:

Embryonic shoot apical meristem

Hm-B:

hygromycin-B

NAA:

naphthaleneacetic acid

SAM:

Shoot apical meristem

2,4-D:

2,4-dichlorophenoxyacetic acid

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Acknowledgements

This work was carried out with financial support from Department of Biotechnology, Government of India.

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  1. Department of Plant Molecular Biology, University of Delhi South Campus, New Delhi, 110 021, India

    P. Baskaran & Indranil Dasgupta

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  1. P. Baskaran
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  2. Indranil Dasgupta
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Correspondence to Indranil Dasgupta.

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Baskaran, P., Dasgupta, I. Gene delivery using microinjection of agrobacterium to embryonic shoot apical meristem of elite indica rice cultivars. J. Plant Biochem. Biotechnol. 21, 268–274 (2012). https://doi.org/10.1007/s13562-011-0078-x

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