The glioma cell lines U87MG or M059K cells were transduced by the packed lentivirus. Shortly, around supplier Lapatinib 2?106 293T cells were seeded in a 100mm dish over night. The lentiviral vector miR 100 or lentiviral vector alone and pPACKH1 Packaging Plasmid Mix were transfected to 293T cells by utilizing LipofectamineTM 2000 in line with the manufacturers instructions. The culture medium containing the packaged infections was collected at 48 h after transfection and was spun at 4 C, 3000rpm for 10 min. The supernatant was collected and polybrene was put into the last concentration 8_g/ml. The mixture was put into the glioma cell culture in a 100mm bowl with 5ml of medium. The transduced cells were harvested after 72?96 h postinfection for further experiments. Cells transfection with 100nM siRNA of PRKDC, ATM, Dicer or hsa miR 100 inhibitor was performed with the lipofectamineTM 2000 according to the manufacturers instructions. Cells were collected at 36 h after transfection for further experiments. The DNA PKcs antibody was purchased from Thermo Cellular differentiation Fisher Scientific Inc.. The ATM antibody and the mTOR antibody were obtained from Cell Signaling. The Ku70 antibody was purchased from Santa Cruz Biotech Inc.. Cycloheximide was purchased from Sigma?Aldrich Inc.. 293T cells were transfected with the appropriate plasmids with or without 100nM hsa miR 100 mimics in 48 well plates. The cells were collected 48 h after transfection, lysed and analyzed with a assay Kit according to the manufacturers protocol and were tested on a luminescence microplate audience LUMIstar Galaxy. order Lenalidomide _Galactosidase or renilla luciferase was employed for normalization. Cell sensitivity to radiation was based on the increasing loss of colony forming ability. Fleetingly, following the cells were irradiated by using an X ray machine at 320 kV, 10 mA, with the filtration of 2 mm aluminum. The dose rate was 2 Gy/min. After IR, the cells were collected and plated, looking at a of 20?100 colonies per plate. Two reproduce meals were prepared for each datum level, and cells were incubated for 14 days to allow cities to develop. Colonies were stained with crystal violet before counting. Statistical analysis of data was done using the Students t test. Differences with p 0. 05 are thought significant. To check for the major reason for the low amount of ATM in M059J cells, we first examined whether there is any difference in ATM during the transcriptional process between M059J and M059K cells by comparing the ATM mRNA levels in both cell lines. The outcome showed that there was no apparent difference in ATM mRNA amounts between M059J and M059K cells. Further real-time RT PCR information confirmed the results, which can be in line with the previous statement, indicating that the low level of ATM in M059J cells is not as a result of low mRNA level.