Maintaining a healthy work–life balance is important for both males and females. Nevertheless, gender segregation still exists in labor markets in South Korea. Therefore, this study aimed to investigate differences in occupational characteristics, job quality indices, and work-life balance between male and female employees. This study was a secondary analysis of the data collected through the fifth Korean Working Conditions Survey in South Korea. Generalized ordinal logistic regression analysis was carried out to investigate the associations between job quality indices and work–life balance of employees by gender. The job quality indices were different according to gender. Male employees were mainly affected by working time quality and work intensity, while female employees were affected by both these factors and by the physical environment. Therefore, strategies differentiated by gender are necessary to improve work–life balance. In particular, more careful attention should be paid to female workers’ physical environment.