Purpose: We aimed to determine gender differences in students' moderate to vigorous physical activity (MVPA) levels measured in primary school by accelerometry. Methods: Our systematic search (PROSPERO: CRD42023422799) was developed using four electronic databases (from January 2013 to December 22, 2023). Our meta-analysis was conducted based on a random-effects model. Results: The search yielded 6255 manuscripts. Only 24 manuscripts were included in the systematic review, reporting 7550 participants. Finally, 19 manuscripts were meta-analyzed. Our analyses indicated that the mean percentages (SMD = 0.32, 95% CI = 0.24 to 0.40, p < 0.001) of MVPA in primary school physical education (PE) lessons were 28.75 ± 13.75 for boys and 25.99 ± 12.35 for girls. In addition, the mean minutes (SMD = 0.22, 95% CI = 0.15 to 0.27, p < 0.001) were 13.75 ± 9.91 for boys and 12.15 ± 8.88 for girls. Students failed to meet the 50% recommendation of MVPA lesson time. Conclusion: Boys spend significantly more time in MVPA than girls during PE lessons in primary school. The findings show the need for methodological changes in PE to reduce this gender gap.