UE Evolution of marine Phytoplankton & Biogeochemistry
Research Worshop 2 :
Functional diversity among Marine Synechococcus
Christophe Six
UMR UPMC-CNRS 7144 “Adaptation & Diversité en Milieu Marin”
Equipe “Procaryotes Photosynthétiques Marins"
Station Biologique de Roscoff
Université Pierre et Marie Curie (Paris VI).
Bureau 354, 3ème étage (Batiment GT)
[email protected]
Research workshop 2
. A scientific approach
. Bringing out a scientific question from a scientific context
. Setting up a experiment plan
. Carring out the experiments
. Synthetising and presenting the results
. Interpreting the results
. Communicating the results and their interpretation properly
. Methods : culturing phytoplankton, flow cytometry and biophysics of photosynthesis,
phylogeny
. Results and interpretation : together
. Personal work : a short report in the form of a research article
 Introduction, Methods, Results, Discussion
 Mini-article due by January 29th on pdf Format at [email protected]
Chlorophyll concentration in the World Ocean
0.1
µg/L
0.5
10
Latitudinal environmental gradients in the World Ocean
0.1
µg/L
0.5
10
?
Latitudinal temperature gradient in the World Ocean
Average temperature for year 2005
Latitudinal
Light
irradiance gradient
in the World Ocean
July 2005
December 2005
Latitudinal environmental gradients in the World Ocean
0.01
1
Chl µg/L)
« Longitudinal » Environmental gradients
In the World Ocean
60
Research Workshop 2
Functional diversity among marine Synechococcus
Global scientific questions :
How Synechococcus cells can adapt to such contrasted environmental conditions ?
Did the evolution shape different cyanobacterial « ecotypes » ?
More focused questions :
.
What may influence the distribution of different pigment types ?
What may inflence the distribution of the different phylogenetic clades ?
What may inflence the pigment cell content ?
What may inflence the photosynthetic activity ?
How to start giving elements of answers to these questions ?
Research Workshop 2
Functional diversity among marine Synechococcus
. To take advantage of the Roscoff Culture Collection
>3000 phytoplankton strains from all aroud the world,
including  350 Synechococcus
Research Workshop 2
Functional diversity among marine Synechococcus
 Panel of 16 Synechococcus strains from the North Atlantic Ocean
 3-4 strains for 2-3 students (triplicate cultures)
 2 sessions to acquire all the data (Monday-Tuesday)
Post 1: Characterise the strains and their isolation sites and build a preliminary phylogeny
 At least one laptop by student triplet
. Gathering information on your strains : Strain denominations, coordinates, depth,
ocean, sea, etc.
Use the RCC website to build a table on an Excel sheet
Install Google Earth (http://www.google.fr/earth/download/ge/agree.html)
. Build a preliminary phylogeny : Retrieve 16S sequences from NCBI and other databases,
align and truncate the sequences on the BIOEDIT software. Then build a neighbor joining
phylogeny with bootstrap values on the MEGA5 software
Install Bioedit (http://www.mbio.ncsu.edu/bioedit/bioedit.html)
 Install MEGA5 (http://www.megasoftware.net/mega.php)
Research Workshop 2
Functional diversity among marine Synechococcus
Post 2: Determine the pigment type (photosynthetic antenna) of each strain by
spectrofluorimetry
 Justine Pittera (PhD student), spectrofluorimeter at the plankton lab
. Determine phycobiliprotein composition
. Calculate phycobilin fluorescence ratios
. Deduce the pigment type
Research Workshop 2
Functional diversity among marine Synechococcus
Post 3 : Determine the chlorophyll cell content for each strain (spectrophotometry)
Determine the cell concentration and the strain cytometry parameters
. Compare the chl a cell content with fluorescence parameters
. Compare to cell size proxy
 Cytometry analyses on Wednesday 16th
Research Workshop 2
Functional diversity among marine Synechococcus
Post 4 :Study photosynthetic capacities by Pulse Amplitude Modulated Fluorimetry
. Using the Junior PAM, draw a fluorescence light response curve
. Determine photosynthetic parameters
Research Workshop 2
Functional diversity among marine Synechococcus
. TD sessions
 Synthetise all the data for all the strains :
- Table with all information
- Whole phylogenic tree with bootstrap values
- All fluorescence spectra with pigment types determined
- Pigment contents and cytometric parameters
- Light response curves
 Compare the different strains
Try to understand whether some features seem to be associated to isolation sites
Considering environmental gradients (latitude, longitude, depth, etc.)