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Open Access Research article

Design and characterization of genetically engineered zebrafish aquaporin-3 mutants highly permeable to the cryoprotectant ethylene glycol

François Chauvigné1, Esther Lubzens2 and Joan Cerdà1*

Author Affiliations

1 Laboratory of Institut de Recerca i Tecnologia Agroalimentàries (IRTA)-Institut de Ciències del Mar, Consejo Superior de Investigaciones Científicas (CSIC), 08003 Barcelona, Spain

2 Department of Marine Biology, Israel Oceanographic and Limnological Research, 81080 Haifa, Israel

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BMC Biotechnology 2011, 11:34  doi:10.1186/1472-6750-11-34

Published: 8 April 2011

Additional files

Additional file 1:

Effect of pH on the permeability of control and DrAqp3b-expressing Xenopus laevis oocytes determined in solutions with diluted or undiluted ion concentrations. Osmotic water permeability (Pf; A) and ethylene glycol permeability (PEG; B) of oocytes expressing 2 ng cRNA of DrAqp3b at different pH. In A, oocytes were preincubated in normal MBS or in isotonic MBS containing 78 mM NaCl and 20 mM sucrose, and subsequently assayed for Pf using 10 times diluted MBS or a hyposmotic bathing solution made by removing the sucrose, respectively. The same values of Pf were obtained by using MBS containing 50 mM or 25 mM NaCl, and 56 mM or 126 mM sucrose, respectively (data not shown). In B, oocytes were preincubated with normal MBS or in isotonic MBS containing 38 mM NaCl and 100 mM sucrose, prior to the swelling assays in isotonic MBS containing 100 mM ethylene glycol. In both A and B, values are the mean ± SEM of a representative experiment (n = 6 oocytes).

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Additional file 2:

Functional characterization of additional DrAqp3b mutants in Xenopus laevis oocytes. Osmotic water permeability (Pf) of oocytes expressing wild-type DrAqp3b (DrAqp3b-WT) or different DrAqp3b mutants at different pH. Values are the mean ± SEM of 2-3 experiments (n = 8-10 oocytes per construct). The asterisks indicate significant differences between DrAqp3b-WT and mutants at a given pH (Student's t test, p < 0.05).

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Additional file 3:

Forward and reverse primers employed to introduce mutations into the zebrafish Aqp3b cDNA. The table lists the oligonucleotide primers employed for the site-directed mutagenesis of the zebrafish Aqp3b cDNA.

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