In order to investigate the effect of clinical application of chicken Mycoplasma synovialis（MS）inactivated vaccine（YBF-MS1strain），the laying rate，hatchability，incidence of respiratory and leg diseases in the offsprings of breeding hens vaccinated with YBF-MS1 vaccine twice and those without vaccination，as well as the reduction rate of laying and shell damage of commercial laying hens were compared and analyzed. The results showed that the laying rate increased by 1.2%-15.8% after the breeding hens were vaccinated with YBF-MS1 inactivated vaccine twice，the hatchability increased by 0-7.3% and the disease incidence of their offsprings decreased by 16.7%-75.0%；the reduction rate of laying was improved by 16.7%-100% and the shell damage reduced by 25.0%-100%. In conclusion，MS infection could be well controlled by YBF-MS1 inactivated vaccine，by which，laying rate and hatching rate were greatly improved，and the incidence rate of commercial generation chickens as well as the incidence of abnormal apical syndrome（EAA）in commercial laying hens were reduced. To this end，future evaluation on clinical application of YBF-MS1 would be supported by the data provided in the study.