In this master's thesis we aimed to develop and optimise a method that would allow us assessment of kin discrimination for bacterium Bacillus subtilis on the cellular level. We concentrated on developing agar pads method to monitor growth and interactions of B. subtilis cells that are genetically more (»kin«) or less (»non-kin«) related by confocal microscopy. We tested the influence of media composition, temperature, time of incubation, inoculation procedure on colony growth. Results did not confirm antagonism between strain, between individual or at the level of microcolonies that we were able to track over time (up to 9 hours) by confocal microscopy. Still both colonies formed a demarcated line and cells mostly did not mix. In the occasions when cells of PS-196 were trapped within the colony of PS-216 we noticed morphological changes of the trapped cells (SMM+ 0,5 % glc). The tested strains still preserved the ability to form visually dramatic boundary at the meeting point of two swarms on agar plates. It is possible that antagonism previously confirmed on agar plates depends on cell density as in microcolonies cells where still at the single cell layer and thus exposed to different conditions than those in swarms on agar paltes. We also examined the influence of relatedness on doubling time. The doubling times of kin strain pairs were more homogeneous in comparison to non-kin strains; however, this conclusion is preliminary as we would need to test higher number of biological replicates to claim this with confidence.
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