Glioblastoma is a highly aggressive brain malignancy with a poor prognosis. Its high intratumor heterogeneity contributes to therapeutic resistance, tumor progression and recurrence. We sequenced 31 loci in 11 patients with glioblastoma (including one patient with samples available from the primary and recurrent tumors) to determine the genetic basis and intratumor heterogeneity of glioblastoma. By analyzing the somatic mutations, known driver genes were identified, including EGFR, PTEN and TP53, and the MUC16 gene exhibited the highest mutation rate in the samples examined. Through an evolutionary analysis of the sequencing results, the EGFR p.L861Q mutation was determined to play a role in the progression from the primary tumor to a relapsing tumor in one patient. We analyzed 1403 genes in blood-derived ctDNA that were previously revealed to play a role in tumorigenesis and the progression of cancer. Somatic mutations identified through ctDNA sequencing that match the results of multipoint exon sequencing in tumor tissues were detected, such as EGFR p.L861Q. These findings provide new insights into the intratumor heterogeneity and evolution of glioblastoma. In addition, ctDNA detection in blood samples represents a convenient method to dynamically identify the genetic changes and new therapeutic targets during the treatment of glioblastoma.