February 1st to 5th 2016
Olhão, Portugal
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Methodology: optimizing protein extraction and separation in Fucus macroalgae to study desiccation-tolerance

Scientific Exhibition
Natural Resources
Wednesday, February 3, 2016 -
18:30 to 20:00

Mota, C.F. 1 Pearson, G.A. 2 Ritter, A. 3 Potin, P. 4 Cavaco, S. 5 Simes, D. 6 Schrama, D. 7 Rodrigues, P. 8 Serrão, E.A. 9

1Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal
2Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal
4SBR
5Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal
6Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal
7Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal
8Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal
9Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal

Shifting between marine and terrestrial habitats, Fucus vesiculosus is a brown intertidal alga that can desiccate rapidly when exposed to air, yet forms dense canopies along European intertidal areas. Desiccation, like temperature and high light, often limits the distribution of intertidal organisms. To identify mechanisms responsible for desiccation tolerance, we searched for proteins differentially expressed during the recovery period following intense desiccation at low tide, when repair and protection mechanisms should be activated.First we selected an optimized method for protein extraction from Fucus vesiculosus, an organism rich in polysaccharides and phenolics, which are known to interfere with protein extraction from other macroalgae and terrestrial plants. Of the four methods tested, two that included phenol extraction and were developed specifically for brown algae, were effective with Fucus adult tips, producing well resolved protein spots in 2-DE. A simpler, commercial protocol for plant tissues, with successive methanol and acetone precipitations, resulted in a viscous yellow extract with few discernible proteins. Another method, developed for macroalgae using RNA extraction columns from a kit, also failed to eliminate contaminants causing viscosity or aggregating proteins. To detect differential expression during recovery from desiccation, purified proteins were separated in 2-DE gels, CBB-stained and the resulting spot patterns analysed. Despite good spot separation and resolution, no significant changes in expression were detected between control and recovery treatments. Our results therefore suggest that constitutive expression is the predominant mode of desiccation protection in intertidal fucoid alga.
keywords: 
protein extraction methods, brown algae, 2-DE, differential protein expression, desiccation

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