Objective To analyze the difference in ground reaction force (GRF) between male and female runners and between left and right limbs using the one-dimensional statistical parametric mapping (SPM). Methods Thirty male and female runners participated the running test on force treadmill at the speed of 12 km/h. GRF of consecutive five steps were recorded. The independent-sample t test and paired-sample t test in the SPM package were used to check the male vs female and right limb vs left limb differences. Results Male and female runners showed inter-limb asymmetry in vertical average loading rate (VALR), and the VALR of male runners was significantly higher than that of female runners in the left limb, while no significant differences were found in the right limb. Male and female runners showed significant differences in the right vertical GRF during push-off and medial-lateral GRF during landing, while left medial-lateral GRF presented significant differences during landing and mid stance. Male runners showed right limb vs left limb significance in the anterior-posterior GRF during push-off, vertical GRF during landing and push-off and medial-lateral GRF during landing, while female runners exhibited right limb vs left limb significance in vertical GRF during landing and push-off. Conclusions This study introduced the application of SPM in statistical analysis of GRF, elaborated the procedures and theory, compared the test of time-series data in SPM and discrete value with traditional statistics, and discussed the difference and features in SPM and SnPM, thus providing references for the application in data analysis of sports biomechanics. Comparison in the male vs female and right limb vs left limb revealed the genders and inter-limb symmetrical differences in the landing, mid-stance and push-off phases during stance. The research findings laid the foundation for future investigation of mechanism in running related injuries and strategy of prevention and treatment.