Antibiotic Disc Assay for Measuring Cell Wall Function in Synechocystis sp. PCC6803
Bio-protocol 2016, 6(24): e2071
Cheregi O, Funk C
This protocol describes how to investigate the integrity of the outer cell wall in thecyanobacterium Synechocystis sp. PCC6803 using antibiotics. It is adapted to the agar diffusion test(Bauer et al., 1966), in which filter paper discs impregnated with specified concentrations of antibioticswere placed on agar plates inoculated with bacteria. The antibiotics we tested, interfering with thebiosynthesis/function of bacterial cell walls, will diffuse into the agar and produce a zone ofcyanobacterial growth inhibition around the disc(s). The size of the inhibition zone reflects the sensitivityof the strain to the action of antibiotics, e.g., a mutation in a protein functioning within the cell wall or itsconstruction would render the mutant strain more sensitive to the respective antibiotic. The method has proven to be useful for phenotyping a mutant of Synechocystis sp. PCC6803 lacking all three genesencoding Deg proteases. Deletion of these ATP-independent serine proteases was shown to haveimpact on the outer cell layers of Synechocystis cells.
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