This meta-analysis was conducted to estimate the magnitude of gender difference in three-dimensional (3-D) mental rotation ability and to investigate how factors related to test administration conditions play a role in varying gender difference effect sizes and threatening validity. Individuals' 3-D mental rotation ability was measured by the Purdue Spatial Visualization Tests: Visualization of Rotations (PSVT:R). We integrated 70 effect sizes of gender differences in mental rotation ability measured by the PSVT:R which were obtained from 40 primary studies. The results indicated that male participants outperformed females on the test (Hedges' "g" = 0.57). The I[superscript 2] statistic indicated 41.7 % of variation in effect sizes reflects real heterogeneity. The moderator analysis indicated that male superiority on spatial ability tasks measured by the PSVT:R is related to the implementation of time limits. The gender difference became larger when stringent time limits (equal or less than 30 s per item) were implemented.