Objective: To investigate the spectrum of gene mutations in adult patients with B-acute lymphoblastic leukemia (B-ALL), and to analyze the influences of different gene mutations on prognosis. Methods: DNA samples from 113 adult B-ALL patients who administered from June 2009 to September 2015 were collected. Target-specific next generation sequencing (NGS) approach was used to analyze the mutations of 112 genes (focused on the specific mutational hotspots) and all putative mutations were compared against multiple databases to calculate the frequency spectrum. The impact of gene mutation on the patients' overall survival (OS) and recurrence free survival (RFS) was analyzed by the putative mutations through Kaplan-Meier, and Cox regression methods. Results: Of the 113 patients, 103 (92.0%) harbored at least one mutation and 29 (25.6%) harbored more than 3 genes mutation. The five most frequently mutated genes in B-ALL are SF1, FAT1, MPL, PTPN11 and NRAS. Gene mutations are different between Ph+ B-ALL and Ph- B-ALL patients. Ph- B-ALL patients with JAK-STAT signal pathway related gene mutation, such as JAK1/JAK2 mutation showed a poor prognosis compared to the patients without mutation (OS: P=0.011, 0.001; RFS: P=0.014,<0.001). Patients with PTPN11 mutation showed better survival than those without mutation, but the difference was not statistically significant (P value > 0.05). Besides, in Ph+ B-ALL patients whose epigenetic modifications related signaling pathway genes were affected, they had a worse prognosis (OS: P=0.038; RFS: P=0.047). Conclusion: Gene mutations are common in adult ALL patients, a variety of signaling pathways are involved. The frequency and spectrum are varied in different types of B-ALL. JAK family gene mutation usually indicates poor prognosis. The co-occurrence of somatic mutations in adult B-ALL patients indicate the genetic complex and instability of adult B-ALL patients.