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After 16 h incubation at 40 °C, cycloheximide was added into medium to inhibit further protein synthesis, after 2 h treatment cells were infected with Legionella for another 2 h then washed 3 times with PBS and cultured with fresh medium at 32 °C to remove the bacteria outside host cells, and then moved to 32 °C for different time points to release VSVG from ER

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