The objective of this study was to evaluate the comparison of production efficiency of oocytes and OPU (ovum pick‐up) derived embryos of Hanwoo with Holstein. The OPU session of each species (6 cows) was carried out from the Hanwoo (106 sessions) and Holstein (114 sessions) at intervals of 3 4 days (2 times per week) for 3 months. Cumulus‐oocyte‐complexes (COCs) retrieved were classified into 4 grades by the status of oocyte cytoplasm and cumulus cells. The COCs collected were matured in vitro in TCM‐199 supplemented with 10% FBS, 10 mg/ml FSH and 1 mg/ml estradiol‐ 17β in 5% CO2 and over 99% humidity for 24 h. After 24 h co‐incubation with post‐thaw sperm, the presumed zygotes were cultured in CR1aa medium with 4 mg/ml BSA for 3 days and then changed CR1aa medium with 10% of FBS for another 3 4 days. The Mean number of aspirated follicles and collected oocytes were not significantly different between Hanwoo and Holstein spacies (10.4±0.42 vs. 11.4±0.41 and 7.5±0.38 vs. 6.1±0.37 per session). But the collection rate of oocytes from aspirated follicles were significantly higher in Hanwoo (72.8%) than that in Holstein (53.6%) (p< 0.05). Furthermore, the average number of good quality oocytes (Grade I and II) was 5.9±0.28 and 4.1±0.27 (Mean±SD), and the cleavage rate and the development rate to blastocysts was significantly (p<0.05) higher in Hanwoo (40.0%) than Holstein (21.6%). The OPU derived embryos of Hanwoo were transferred 83 times into 52 recipients and then 42 calves were produced from 44 pregnancy recipients. In conclusion, the efficiency of OPU derived embryo was significantly different between Hanwoo and Holstein species. In vitro culture system for OPU derived embryo production should be optimized for industrialization and the improvement of livestock.