Abstract
Cyanobacteria are prokaryotic organisms performing oxygenic photosynthesis. The cyanobacterium Synechocystis sp. PCC 6803 is a model organism for the study of photosynthesis, gene regulation and biotechnological applications because it is easy to manipulate genetically. Moreover, this cyanobacterium can grow photoautotrophically as well as chemoheterotrophically in the dark utilizing glucose. Microbiologists often use optical density measured with a spectrophotometer for the comparison of growth performance of different strains in liquid cultures. Because Synechocystis sp. PCC 6803 (especially motile strains) tend to form aggregates under stress conditions this method might be not suitable for evaluation of different strains under different growth conditions. In addition, many labs are not well equipped with standardized photobioreactors and illumination facilities to ensure reproducibility of growth curves. Here, we describe a highly reproducible spot assay for viability analysis of Cyanobacterial strains.
Keywords: Cyanobacteria, Synechocystis 6803, Spot assay, Growth assay
Materials and Reagents
Equipment
Procedure
Every experimental step involving the handling of cyanobacterial cell cultures must be performed under a Laminar flow hood to prevent contamination!
Notes
The spot assays described above are highly reproducible. In contrast to other methods like measuring the optical density of liquid cultures, which is often error-prone due to i.e. the formation of aggregates, we always obtained reliable and reproducible results. Within our group, several researchers have performed this method in two different laboratories and we did not detect any variations in the results. For reproducibility it is highly recommended to use Na2S2O3 in the plates (as described in Recipes) and to record the results after the same time. Composition of agar plates and light conditions can be customized to fit the experimenters’ individual needs and may require optimization for different mutant strains.
Recipes
Acknowledgments
BG11 medium is prepared according to Rippka et al. (1979). This work was supported by DFG grant to A. W. Wi2014/5-1.
References
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