Objective To explore the correlation of fatigue with health-promoting lifestyle in the udnergraduate students, so as to provide scientific evidence for the intervention. Methods A cross-section trial was carried out in the undergraduate students from a medical university in Guangzhou. By a self-designed questionnaire, the incidence of fatigue in the undergraduate students was figured out after data processing and statistical analysis. Results A total of 6 100 questionnaires were released, and 5 843 were responded, with a respondent rate being 95.79%. The number of effective questionnaires arrived 5 676. The results showed that the prevalence of fatigue among undergraduate students was 49.6%. The female had a higher risk than the male ( 51.6% vs 45.8%, P<0.01). The prevalence of fatigue in grade 1-3 students was 49.4%, 48.1% and 52.1% , respectively, the difference being insignificant among the three grades (P>0.05). The students with poor level of health-promoting lifestyle had a higher risk of suffering from fatigue (OR=6.647, 95%CI: 4.913-8.993). Conclusion Fatigue is prevalent among the undergraduate students. The incidence of fatigue can be reduced by increasing the level of self-actualization, physical exercise and pressure management.