Background: Astrocytoma and glioblastoma (GBM) are two main subtypes of glioma classified by WHO guide of center nervous system (CNS) tumors to different grades. GBM is the most malignant among all CNS tumors with a 5-year survival rate of less than 5%. Although the prognosis of patients with astrocytoma is better than that of GBM in general, astrocytoma patients with IDH wild-type have a similar prognosis as GBM. Exploring the molecular driving force behind the malignant phenotype of astrocytoma and GBM will help explain the diversity of glioma and discover new drug targets. Methods: We enrolled 12 patients with astrocytoma and 12 patients with GBM and performed whole-exome sequencing (WES) and RNA-seq analysis on tumor samples from the patients. Comparative analysis of somatic mutation, copy number variation, cytoband alteration and differently expressed genes were performed between astrocytoma and GBM. Immune cells infiltration was analyzed by CIBERSORT Algorithm with LM22 immune subsets.Results: This study includes 14 male patients and 10 female patients, and the median age was 45.5 (range, 18-68) years old. Seven patients had IDH1 mutation, including 6 patients with astrocytoma and 1 patient with GBM. Somatic mutation of KRT18, which is associated with cell apoptosis and adhesion by interacting with TRADD and Pinin, was significantly enriched in astrocytoma, but rare in GBM. Copy number loss of MTAP, which is a potential drug target and closely related to a poor prognosis of glioma, was found significantly enriched in GBM. Pathway enrichment analysis found that GBM was mainly enriched in tumor-related pathways, such as cell adhesion, cell division, and angiogenesis, but astrocytoma was mainly enriched in pathways with ion transport, cell-cell signaling and neurogenesis. Immune cells infiltration analysis showed that macrophages M0 was significantly enriched in GBM, and activated mast cells, monocytes and plasma cells were significantly enriched in astrocytoma. Conclusions: This study revealed the distinct characteristics of astrocytoma and GBM at the DNA and RNA levels. Somatic mutation of KRT18, which was first reported in glioma, may be closely related to the malignant phenotype of astrocytoma. Copy number loss of MTAP, which was associated with the susceptibility to purine starvation and specific chemotherapy, was a key genetic alterative gene in GBM and had the potential to become the therapeutic target of GBM.