Small RNA sequencing for Ad_24h
Human primary lung fibroblast cells (IMR-90) (American Type Culture Collection) were cultured in Eagle's minimum essential medium (MEM) with Earle's salt and GlutaMAX™ supplemented with 10% fetal bovine serum, 100 U/ml penicillin, 100 µg/ml streptomycin, 0.1 mM nonessential amino acids, 1.5 g/L sodium bicarbonate and 1.0 mM sodium pyruvate. After reaching confluence, the cells were cultured for two more days in order to synchronize the cells. Over 95% of the cells were in the G0/G1 phase as indicated by FACS analysis Synchronized monolayer cells were mock-infected or infected with Ad2 at a multiplicity of 100 fluorescence-forming units (FFU) per cell in serum-free medium. After 1 h adsorption at 37 °C, the medium was replaced with MEM containing 10% FBS and incubated at 37 °C. Infected cells were collected at 6, 12, 24, and 36 h post infection (hpi). Mock-infected cells were collected at 6 hpi.