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A new short term toxicity assay using Aspergillus awamori with recombinant aequorin gene.

Kozlova, Olga, Zwindermann, Mark and Christofi, Nicholas (2005) A new short term toxicity assay using Aspergillus awamori with recombinant aequorin gene. BMC Microbiology, 5 (1). pp. 1-9. ISSN 1471 2180

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    Abstract/Description

    Background
    Most currently available short-term toxicity assays are based on bacterial cells. Therefore there is a need for novel eukaryotic microbial bioassays that will be relevant to higher eukaryotes such as animals and plants. Ca2+ is a universal intracellular signalling molecule found in all organisms from prokaryotes to highly specialized animal cells. In fungi calcium has been demonstrated to be involved in control of many important processes. The recombinant aequorin gene from the jellyfish Aequorea victoria responsible for the expression of the Ca2+-sensitive aequorin photoprotein has been cloned in the filamentous fungus Aspergillus awamori. This has allowed real life monitoring of [Ca2+]c changes in living fungal cells. When subjected to different physico-chemical stimuli fungal cells respond by transiently changing the concentration of free Ca2+ in the cytosol ([Ca2+]c) and the pattern of these changes (Ca2+ signature) is specific to each particular stimulus. Therefore it was interesting to investigate whether different environmental toxicants would be able to affect the pattern of [Ca2+]c changes in a reproducible and dose dependant manner.


    Results
    Toxicity bioassay has been developed to monitor changes [Ca2+]c of the recombinant fungus in the presence of toxicants representing heavy metals – Cr6+ and Zn2+ and a phenolic polar narcotic -3,5-DCP. The fungus responds to toxicants by a decrease in the amplitude of [Ca2+]c response to 5 mM external CaCl2 and an increase in Ca2+ final resting levels and recovery time.


    Conclusion
    A novel toxicity bioassay utilizing eukaryotic cells has been developed based on filamentous fungi transformed with the recombinant aequorin gene. A range of parameters characterising changes in [Ca2+]c has been identified, e.g. Amplitude, Length of Transient, Final Resting Level and Recovery Time. These parameters can be used to determine the toxicity of a range of chemicals to eukaryotic cells in a 96-well microtitre plate method.

    Item Type: Article
    Print ISSN: 1471 2180
    Additional Information: PMCID: PMC1177953 is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
    University Divisions/Research Centres: Faculty of Health, Life & Social Sciences > School of Life Sciences
    Dewey Decimal Subjects: 500 Science > 570 Life sciences; biology > 579 Microorganisms, fungi & algae
    300 Social sciences > 360 Social problems & social services > 363 Other social problems & services > 363.7 Environmental pollution
    Library of Congress Subjects: G Geography. Anthropology. Recreation > GE Environmental Sciences
    Q Science > QR Microbiology
    Item ID: 1718
    Depositing User: RAE Import
    Date Deposited: 14 Jul 2008 13:06
    Last Modified: 16 Apr 2013 16:19
    URI: http://researchrepository.napier.ac.uk/id/eprint/1718

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