Isolation and Cultivation of Plant-Associated Bacteria

To isolate cultivable bacteria, surface sterilized root and shoot tissue of three plants were separated and transferred to mortars with 5 ml sterile 10 mM MgSO₄ to compose a mixed sample. The plant tissues were separately crushed and serial dilutions were prepared (0, 10⁻¹, 10⁻², 10⁻³, 10⁻⁴). One hundred microlitre of each dilution was applied to the Petri dishes containing different media (Table 1); all Petri dishes were established in triplicate. The Petri dishes were incubated at 30°C for 4 days after which the cfu per gram of fresh plant tissue was determined; the averages and standard errors were calculated for each treatment. The colonies were then purified and 584 strains were stored at −80°C in glycerol (15%_w glycerol, 0.85%_w NaCl).