Distribution and composition of microplankton

1
Title
Distribution and composition of microplankton
along the SE Pacific Ocean
Fernando Gómez1 Hervé Claustre2
1Department of Aquatic Biosciences, The
University of Tokyo
2Laboratoire dOcéanographie de
Villefranche-sur-Mer
address until February 2004
fernando.gomez_at_fitoplancton.com
2
Objectives
To study the distribution (vertical and
longitudinal) and composition of microplankton
from the eutrophic (coastal upwelling of Chile)
to ultra-oligotrophic regimes (South Pacific
gyre). To study the temporal variations on the
distribution (diel cycle) in selected stations
To investigate the occurrence and distribution of
N2-fixers (Trichodesmium spp.) and N2-fixer
symbiotic associations (Richelia and others)
To contribute to the study of marine
phytoplankton taxonomy and biodiversity in one of
the less studied major oceanic entities of the
world ocean
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Sample collection
Niskin bottle
3 ml Lugol
station, depth
500 ml
seawater
Polyestyrene plastic bottle
Storage dark, cool and quiet place
0, 5, 10, 20, 30, 50, 70, 90, 100, 120, 150, 175,
200 m depth
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Methodology
Methodo
Pre-concentration
500 ml Lugol fixed sea-water samples
Settling in counting chambers
Microscopical analysis (inverted microscope)
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Taxonomical studies 1. Light microscopy
The specimens of interest will be isolated with a
capillary from the chambers
High magnification microphotographs (1000)
Nomarski Differential Interference Contrast
(D.I.C.)
Morphology and location of organelles such as
chloroplasts, pyrenoids, nuclei, cellulose
thecal plates, flagella, etc
transferred to a glass slide
staining compounds (flourochromes DAPI,
Fluorescent Brightener, etc )
Fluorescence microscopy (UV light or blue light
according to the staining compound)
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Taxonomical studies 2. SEM
Scanning Electron Microscopy
The specimens of interest will be isolated with a
capillary from the chambers
Takayama method
Adhered on the poly-L-lysine coated glass plate
rinse in distilled water dehydrate through an
ethanol series
Ion spatter coating with Au or Au-Pd
Scanning Electron Microscopy
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Taxonomical studies 3. Gene sequencing
After to complete the morphological
characterization of the species of interest, the
forthcoming specimens will be picked and keep
for single-cell molecular analysis (phylogeny)
Discrepancies on the most appropriate fixative
(methanol) for Polymerase Chain Reaction (PCR)
appear in the literature
The protocol of extraction is quite similar, the
gene amplification protocol varies according to
the instruments and reagents suppliers

Nucleotide sequences of coding (small subunit
(SSU), partial large subunit (LSU) (specific
diversity) and internal transcribed spacer
region (ITS1-5.8SSU-ITS2) parts of the rRNA
operon (intraspecific diversity)