To investigate the relationship between DNA-PKcs and genome instability & hyper-radiosensitivity, human glioma cell lines M059K and M059J, as a model expressing wild-type DNA-PKcs and a model defective in DNA- PKcs activity, were exposed to low doses of X-rays. Cells survival fractions were assessed by colony-forming assay and Cytochalasin-B micronucleus assay was employed to detect the genomic instability happening in each single irradiated colony. It has been found that as the post-incubation time increased, M059K cells expressing wild-type DNA-PKcs exhibited low-dose hyper-radiosensitivity and showed a similar genomic instability after 0.2 Gy and 0.6 Gy irradiations, but the M059J cells lacking in DNA-PKcs didn’t present low-dose hyper-radiosensitivity and showed a higher genomic instability of 0.6 Gy than that of 0.2 Gy. The results indicate that DNA-PKcs may act as one of the key factors that lead to low-dose hyper-radiosensitivity.