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Microscopy analysis
Stereo microscope
Stereo microscope
The stereo microscope was used to observe terrestrial algae, which was sampled from university building and Trentepohlia umbrina was separated from its subsurface to upset a microalgae culture.
Inverse microscope
Inverse microscope
In comparison to terrestrial algae, aquatic species were mainly analyzed applying an inverse microscope. These microscopes are optimal to investigate liquid samples, containing microalgae. Besides identification and quantitative analysis of marine and freshwater species, this technique enables the isolation of single cells which are used to create monoclonal cultures.
Scanning Electron Microscope (SEM)
Scanning Electron Microscope (SEM)
The SEM was applied to visualize the morphology of Pseudo-nitzschia multistratia shells, which are cleaned before with acid. Due to the high magnification up to 60Kx of the electron microscope and the data processing using SMARTSEM software, details like the stria pattern and the raphe are visible. Two different lenses are used to capture the electrons which are diverted from the sample. The InLens is directly located near the electron emitter while the position of the 2nd lens receives the electrons at a greater distance.
Physical analysis
Autofluorescence measurements using FlowSight were performed to investigate the metabolic states of Chaetoceros muelleri cells, which have been previously exposed to thermal stress for different durations. The measurements were carried out as a preliminary test to measure the resistance of this species to different degrees of stress (Surre et al., 2018).
FlowSight autofluorescence gauge
Chemical solutions
Preparation of nutrient medium
F2 medium for diatoms was prepared using 2 mL NaNO3, 2 mL NaH2PO4, 2 mL Na2SiO3, 2 mL HEPES, 2 mL trace metals, and 1 mL vitamin solution in 2 L of autoclaved and filtered seawater with a salinity of 30 ppt.
For F4 medium, to cultivate dinoflagellates, 1 mL NaNO3, 1 mL NaH2PO4, 20 µL Na2SeO3, 2 mL HEPES, 2 mL trace metals, and 1 mL vitamin solution was added to 2 L seawater with a salinity of 35 ppt.
Trace metals solution contains essential metals for the primary production. 3.15 g FeCl3, 4.36 g Na2EDTA, 1 mL MnCl2, 1 mL ZnSO4, 1 mLCoCl2, 1 mL CuSO4, and 1 mL Na2MoO4 are added to 995 mL distilled and autoclaved water.
HEPES stock is prepared, dissolving 23 g (HEPES buffer) in 100 mL autoclaved and distilled water.
To prepare the vitamin solution, 0,1 g biotin (vitamin H) was dissolved into 100 mL distilled water and 0,1 g Cyanocobalamin (vitamin B12) in 100 mL in another bottle. 200 mg Thiamin is directly dissolved in 950 mL of distilled and autoclaved water and 1 mL of each stock is added.
NaH2PO4 stock, contains 0.375 g NaH2PO4, which is dissolved in 75 mL distilled and autoclaved water.
(Andersen, 2005)
Genetic analysis
During the internship, I extracted DNA from four diatoms cultures of Pseudo-nitzschia multistratia and one benthic unknown diatom culture using CTAB protocols. After replicating the DNA with PCR, agarose gel is prepared and the electrophoresis is started. The analysis of the gel did not yield any significant results so that further genetic analyses will be continued in the future.
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