Abstract:
The present study was designed to assess the similarity of follicular development, oocyte quality, and their subsequent development on ovum pick-up (OPU)-IVF in identical twin cattle. Four pairs of identical twin Japanese black cows (A, B pairs at 5 years old and C, D pairs at 3 years old) were kept under the same feeding and environmental conditions. OPU was performed for these cows once a week for seven continuous weeks. OPU was done by using a 7.5-MHz linear transducer with needle (17 G, 530-mm length) connected to an ultrasound scanner (SSD-1200, ALOKA, Tokyo, Japan). Oocytes were evaluated by their cumulus cell morphology, cytoplasmic color, and density. To assess the development, collected COCs were cultured for 20�h in TCM-199 supplemented with 5% calf serum (CS) in a microdroplet (volume was adjusted to 5�_L/oocyte) at 38.5�C under atmosphere of 5% CO2 in air. After maturation, the COCs were inseminated with frozen-thawed semen collected from the same ejaculation of a single bull. The fertilization was performed with BO solution as described by Imai et al (J. Vet. Med. Sci., 2002, 64(10), 887-891). The zygotes were then cultured in CR1aa supplemented with 5% CS under the same condition of maturation for nine days. Embryo development was assessed by the cleavage rate on Day 2 and the blastocyst production rate on Days 7 to 9 (insemination day�=�Day 0). Blastocysts were classified according to the IETS creteria. Data were analyzed by ANOVA. A total 56 sessions of OPU were performed in this study. The overall mean number of developing follicles (larger than 2�mm in diameter), collected oocytes, and produced blastocysts were 30.3ʱ�9.2, 20.1ʱ�9.2 and 6.3ʱ�3.8 (meanʱ�SD) per session, respectively. The mean number of developing follicles on the day of OPU were significantly different between B and D pairs (38.6ʱ�7.5 and 21.9ʱ�6.5, P�<�0.01); however, no significant difference was found within each twin. In oocyte quality, C and D pairs were significantly higher grade than the A pair. The percentages of cleaved oocytes and embryos developed to the blastocyst stage (34ʱ�16, 27ʱ�10, 41ʱ�17 and 39ʱ�24) showed no differences among 4 pairs and within each twin. However, the percentage of Grade 1 blastocyst of B pair was significantly lower (P�<�0.01) than that of other pairs, and C pair was significantly higher (P�<�0.05) than that of A and D pairs (67ʱ�25, 41ʱ�22, 93ʱ�10 and 71ʱ�25; A, B, C and D pairs, respectively). There was no significant difference within twins. These results show little statistical variation between cows of the same genetic background in the production of embryos in vitro.