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Expression of recombinant Atlantic salmon serum C-type lectin in Drosophila melanogaster Schneider 2 cells

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Abstract

The Atlantic salmon (Salmo salar) serum lectin (SSL) is a soluble C-type lectin that binds bacteria, including salmon pathogens. This lectin is a cysteine-rich oligomeric protein. Consequently, a Drosophila melanogaster expression system was evaluated for use in expressing SSL. A cDNA encoding SSL was cloned into a vector designed to express it as a fusion protein with a hexahistidine tag, under the control of the Drosophila methallothionein promoter. The resulting construct was stably transfected into Drosophila S2 cells. After CdCl2 induction, transfected S2 cells secreted recombinant SSL into the cell culture medium. A cell line derived from stably transformed polyclonal cell populations expressing SSL was used for large-scale expression of SSL. Recombinant SSL was purified from the culture medium using a two-step purification scheme involving affinity binding to yeast cells and metal-affinity chromatography. Although yields of SSL were very low, correct folding and functionality of the recombinant SSL purified in this manner was demonstrated by its ability to bind to Aeromonas salmonicida. Therefore, Drosophila S2 cells may be an ideal system for the production of SSL if yields can be increased.

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Acknowledgments

We thank Sandra Sperker and Robert Richards for technical advice and we thank Doug Kuntz and Dr. Santosh Lall for review of the manuscript. We are grateful to Dr. Catherine Too for loan of the tissue culture flasks. This study was supported by an Izaak Walton Killam Predoctoral Scholarship (E. Uribe), an NSERC Discovery grant (K. V. Ewart) and by the NRC Institute for Marine Biosciences. This is NRC publication number 54062.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Dalhousie University, Halifax, NS, B3H 1X5, Canada

    Elke Uribe

  2. Ontario Cancer Institute, University Health Network, 101 College Street, Toronto, ON, M5G 1L7, Canada

    Meenakshi Venkatesan

  3. Department of Biology, University of Waterloo, Waterloo, ON, N2L 3G1, Canada

    David R. Rose

  4. National Research Council of Canada, Halifax, NS, B3H 3Z1, Canada

    Kathryn Vanya Ewart

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  1. Elke Uribe
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  2. Meenakshi Venkatesan
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  3. David R. Rose
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  4. Kathryn Vanya Ewart
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Correspondence to Kathryn Vanya Ewart.

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Uribe, E., Venkatesan, M., Rose, D.R. et al. Expression of recombinant Atlantic salmon serum C-type lectin in Drosophila melanogaster Schneider 2 cells. Cytotechnology 65, 513–521 (2013). https://doi.org/10.1007/s10616-012-9505-7

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  • DOI: https://doi.org/10.1007/s10616-012-9505-7

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