GSM5061429: Control 1; Rattus norvegicus; RNA-Seq
A CLINIC600C linear accelerator (Varian, United States) was used to irradiate BMECs, at a rate of 2.5 Gy/min. Cells cultured to 60% confluence were irradiated with 5 Gy or 20 Gy. The x-ray source was 100 cm from the cells and had an irradiation range of 20 cm × 20 cm. The cells in the control group were not irradiated by x-rays, but received all other treatments that the x-ray–exposed group received. The primary cultured BMECs were divided into five groups: (1) unexposed controls; (2) 12 h after delivery of 5 Gy of radiation; (3) 24 h after delivery of 5 Gy of radiation; (4) 12 h after delivery of 20 Gy of radiation; and (5) 24 h after delivery of 20 Gy of radiation. RNA was extracted using a commercially available kit (Vazyme Biotech Co., Ltd, Nanjing, China). BMECs with or without radiation exposure were digested and resuspended in lysis buffer RL1. The cell suspension was transferred to a genomic DNA filter column and centrifuged at 13000 × g for 2 min. The resulting supernatant was collected and buffer RL2 was added. The mixture was placed in an RNAPure column and centrifuged again. The precipitate was collected. Buffer RW1 and buffer RW2 were used to rinse the RNAPure columns. The columns were put into new RNase-free collection tubes. RNAase-free ddH2O (60 μL) was added to the central part of the columns. The columns remained at room temperature for 2 min to dissolve RNA, and were then centrifuged at 13000 × g for 1 min to elute RNA. A spectrophotometer was used to determine the purity and concentration of the extracted RNA. The optical density ratio of 260 nm to 280 nm was between 1.8 and 2.2, which met experimental requirements.