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Development of a fluorescent transgenic zebrafish biosensor for sensing aquatic heavy metal pollution

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Abstract

We report a transgenic zebrafish (Danio rerio) designed to respond to heavy metals using a metal-responsive promoter linked to a fluorescent reporter gene (DsRed2). The metallothionein MT-Ia1 promoter containing metal-responsive elements was derived from the Asian green mussel, Perna viridis. The promoter is known to be induced by a broad spectrum of heavy metals. The promoter-reporter cassette cloned into the Tol2 transposon vector was microinjected into zebrafish embryos that were then reared to maturity. A transgene integration rate of 28 % was observed. The confirmed transgenics were mated with wild-type counterparts, and pools of F1 embryos were exposed to sub-lethal doses of Cd2+, Cu2+, Hg2+, Pb2+ and Zn2+. The red fluorescence response of zebrafish embryos was observed 8 h post- exposure to these sub-lethal doses of heavy metals using a fluorescence microscope. Reporter expression estimated by real-time PCR revealed eightfold, sixfold and twofold increase on exposure to highest concentrations of Hg2+, Cd2+ and Cu2+, while Pb2+ and Zn2+ had no effect. This biosensor could be a first-level screening method for confirming aquatic heavy metal bio-toxicity to eukaryotes.

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Acknowledgments

The authors acknowledge the Department of Biotechnology, Government of India and the Indian Council of Agricultural Research, New Delhi for funding this research project; Dr. Dilip Kumar and Dr. W. S. Lakra, former Directors, ICAR-CIFE, Mumbai and Dr. Gopal Krishna, Director, ICAR-CIFE, Mumbai for providing facilities.

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Authors and Affiliations

  1. Department of Fish Genetics and Biotechnology, ICAR-Central Institute of Fisheries Education, Versova, Mumbai, 400 061, India

    Nilambari Pawar, P. Gireesh-Babu, Supriya Sabnis, Kiran Rasal, Renuka Murthy, S. G. S. Zaidi & Aparna Chaudhari

  2. CSIR-Institute of Genomics and Integrative Biology, Mathura Road, New Delhi, India

    Sridhar Sivasubbu

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  1. Nilambari Pawar
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  2. P. Gireesh-Babu
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  3. Supriya Sabnis
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  5. Renuka Murthy
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  6. S. G. S. Zaidi
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  7. Sridhar Sivasubbu
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  8. Aparna Chaudhari
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Correspondence to Aparna Chaudhari.

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Pawar, N., Gireesh-Babu, P., Sabnis, S. et al. Development of a fluorescent transgenic zebrafish biosensor for sensing aquatic heavy metal pollution. Transgenic Res 25, 617–627 (2016). https://doi.org/10.1007/s11248-016-9959-z

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  • DOI: https://doi.org/10.1007/s11248-016-9959-z

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