Development of embryo using bovine and ovine frozen -thawed and freeze-dried spermatozoa injected into bovine oocytes
2007
Koyama, H.(Rakuno Gakuen Univ., Ebetsu, Hokkaido (Japan)) | Hoshino, M. | Tsuchiya, K. | Abe, H. | Sugulle, A.H. | Dochi, O.
The objectives of the present study were to investigate the efficiency of bovine and ovine frozen thawed and freeze-dried spermatozoa injected (ICSI) into bovine oocytes and subsequent embryo development. The semen was collected from three different holstein bulls (B-1, B-2 and B-3) and two ovine sperm (S-1 and S-2). For successful blastocysts development, normal sperm DNA is essential. To determine whether sperm DNA is damaged during freeze-drying, comet assay was performed both bovine and ovine sperm (frozen and freeze-dried). There were no significant differences in normal sperm DNA between the three bulls, both frozen thawed and freeze-dried spermatozoa. Also there were no significant difference in normal sperm DNA between the two ovine, both frozen and freeze-dried sperm. To investigate embryo development (cleavage rate and blastocyst), frozen and freeze-dried bovine spermatozoa were injected into bovine oocytes. In respective of bull, cleavage rate and blastocyst rate were significantly lower than that of control (P0.01). But, there were no significant difference between the three bulls. Frozen and freeze-dried ovine spermatozoa were injected into bovine oocytes, and evaluated the fertilization ability. There were no significant difference between frozen-thawed and freeze-dried ovine sperm after ICSI both number of successfully injected oocytes and number of pronuclear formation. In this experiment, the blastocysts were obtained from the frozen thawed bovine spermatozoa, but not freeze-dried spermatozoa. Pronuclear formation was observed ovine spermatozoa. Both the cleavage rate and the pronuclear formation were very low. So, further research on the injection method and activation of oocytes after injection are needed.
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