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Transmission electron microscopy

SS Simon Sretenovic
BS Biljana Stojković
ID Iztok Dogsa
RK Rok Kostanjšek
IP Igor Poberaj
DS David Stopar
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For negative staining transmission electron microscopy (TEM), ~ 20 μl of samples were transferred and left for 5 min on grids covered with Formvar support film (SPI supplies, USA) and negatively stained with 1 % (w/v) aqueous uranyl acetate for 5 s. B. subtilis and E. coli were grown in SM (SYM44 without yeast extract) and M9 medium (6.4 % (w/v) Na2HPO4 × 7H2O, 1.5 % (w/v) KH2PO4. 0.25 % (w/v) NaCl, 0.5 % (w/v) NH4Cl, 2 mM MgSO4, 0.4 % (w/v) glucose, 0.1 mM CaCl2), respectively, to reduce the polymeric background otherwise present in SYM and LB media. The samples were fixed immediately to prevent further modifications. Excess staining solution was removed by a filter paper. Stained bacteria were air dried at room temperature and examined with a Philips CM 100 electron microscope at 80 keV.

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