"Sampling protocol for population genetics" Method

mardi 2 juin 2015
par  Dorian GUILLEMAIN, Romain DAVID
popularité : 12%

Index of "Sampling protocol for population genetics?"
1. Principle
2. Method
3. Sampling
4. Deliverables
5. To summarize
Back to protocols Index
How to cite this content :

David R., Arvanitidis C., Çinar M.E., Dubar J., Dubois S., Erga Z., Guillemain D., Sartoretto S., Thierry de Ville d’Avray L., Zuberer F., Chenuil A., Féral J.-P., (2014), with contributors : Açik Çinar S., Andral B., Aurelle D., Aysel V., Bakir K., Bellan G., Bellan-Santini D., Bouchoucha M., Celik C., Chatzigeorgiou G., Chatzinikolaou E., Chenesseau S., Dağli E., Dailianis T., Dimitriadis C., D’Iribarne C., Doğan A., Dounas C., Egea E., Elguerrabi W., Emery E., Evcen A., Faulwetter S., Gatti G., Gerovasileiou V., Güçver S.M., Issaris Y., Katağan T., Keklikoglou K., Kirkim F., Koçak F., Koutsoubas D., Marschal C., Önen M., Önen S., Öztürk B., Panayiotidis P., Pavloudi C., Pergent G., Pergent-Martini C., Poursanidis D., Ravel C., Reizopoulou S., Rocher C., Ruiton S., Sakher S., Salomidi M., Sarropoulou E., Selva M., Sini M., Sourbes L., Simboura N., Taşkin E., Vacelet J., Valavanis V., Vasileiadou A., Verlaque M. Protocols for monitoring of coralligenous habitats of mediterannean (Coralligenous based Indicators to Evaluate and Monitor the "good ecological status" of the MEDiterranean coastal waters) Protocoles de suivi du coralligène en méditerranée (Coralligenous
based Indicators to Evaluate and Monitor the "good ecological status" of the MEDiterranean coastal waters).

[§ 1] Sampling is equally performed on different ecological profiles of each site? in order to know the structure of the population according to the profile? and, thus, species ecology.

[§ 2] Since the frequencies of different genetic variants will be compared, it is important to have enough samples to allow statistical analysis on the possible occurrence of significant differences among the frequencies. About thirty samples for each taxa (from thirty different colonies and thalli) should be collected along each transect? (Porter et al., 2002 ; Schwaninger, 1999).

[§ 3] The sampling should be on two sides of the study site, if possible opposites to another and at depth of 28 ±1 m, in a way that the collected samples would come from all possible orientations. In each side, we choose two different profiles (in terms of the inclination and the rugosity of the sustrate) and the majority of segments should be where the studied species are found. These profiles would have been firstly determined by the results of the cartography.

[§ 4] The collected colonies must be separated by about 1 m of each other to not collected the same one several times. They must tie in an ecologic profile well defined and listed at the time of collection. Therefore the numerous of the segment in which the sample is collected has to be in the name of each individual (Figure 1).

[§ 5] Ideally, a minimum of 8 samples of each species should be collected in every one of the two pre-selected profiles (for two well distingued orientations and at two depths), in a way to provide replicas for the analysis. For this reason, the sampling would be in the 5 m segments (predefined by the cartography for the principal studied sites of Marseilles region, or chosen in relavence to their conformity for the secondary sites or for the localities where it wasn’t priory possible to perform the cartography). In every segment where the sampling is possible, a fragment of each species should be collected at a minimum distance of 1 m so that a good repartition of the samples would be achieved.

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Figure 1 : sampling method for M. truncata and L. stictaeforme/cabiochiae.

[§ 6] Ideally, samples should also be collected at wider scale : at least two other regions in order to be able to analyse the correlation between the genetic and the spatial distance. This would permit to deduce a « distance of dispersion within the generations » :
- From the Western Mediterranean Sea, with example some study sites between Nice and Montpellier, but should also make an object of prospecting at out neighbors Italians and Spanish.
- From the Eastern basin, which would be possible with the collaboration of our CIGESMED partners from Greece and Turkey.

Identification of Lithophyllum stictaeforme/cabiochiae

At first, it is necessary to differenciate L. stictaeforme/cabiochiae from Mesophyllum spp. (Figure 2).

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Figure 2 : (1) Mesophullum alternans(and Peyssonnelia squamaria), (2) Lithophyllum stictaeforme, (3) Lithophyllum cabiochiae, (4)Neogoniolithon mamillosum, (5) Peysonnelia rosa-marina (Ballesteros, 2006)

[§ 7] For the identification of species from the inferior surface of the thallus :
-  Inferior surface smooth and whitish -> Mesophyllum spp. (Figure 3(1)).
-  Inferior surface pink, rather rough and finely white striped, often with tubers -> Lithophyllum stictaeforme/cabiochiae (Figure 3(4)).
-  Concentric stripes -> Lithophyllum stictaeforme.
-  Radial and concentric stripes -> Lithophyllum cabiochiae.
For the identification of species from the upper surface of the thallus :
- Pink-orangish upper face, white border -> Mesophyllum spp. (Figure 3(1)).
- Pink-purple rather mat and without white border -> Lithophyllum stictaeforme/cabiochiae (Figure 3(3)).

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Figure 3 : External differences between Mesophyllum spp. and L. stictaeforme/cabiochiae.

[§ 8]In case the morphological identification does not provide satisfying answers, it is necessary to analyse the cellular structure of the samples, through the microscopic observation (stereo microscope) of vertical radial sections (i.e. sections obtained following in the growth pattern of the calcareous blades). The species of the complex L. stictaeforme/cabiochiae show a layer of basal cells (hypothallus) which generate vertical cellular lines (perithallus) slightly sloping (L. stictaeforme) or slightly sloping in the centre of the blades and strongly sloping near the edges (L. cabiochiae). Perithallus cells are never aligned (unlike Mesophyllum spp.).
Mesophyllum spp. cells are ranged concentrically with locally cells fusions (Figure 4(1, 2), while L. stictaeforme/cabiochiae cells (Figure 4(3, 4) have rounded lateral walls and communicate laterally only by secondary synapsis.

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Figure 4 : Radial vertical sections of Mesophyllum expansum (1 and 2) andLithophyllum stictaeforme /cabiochiae (3 and 4) thalli.

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- Deadline : 14th December 2015
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