NUMAN AKYOL, SEDAT HAMDİ KIZIL, Muharrem SATILMIŞ, TAHİR KARAŞAHİN

Investigation of bull effect on in vitro embryo production

Dokuz farklı Holştayn boğaya ait spermanın kullanıldığı bu in vitro embriyo üretim çalışmasının amacı, hem boğalar arasında hem de boğaların kendi tekrarları arasında yarıklanma ve blastosiste ulaşma oranları bakımından fark olup olmadığının gösterilmesi olmuştur. Suni tohumlama boğası olarak kullanılan ve fertilite sorunu olmayan ve tatminkar düzeyde fertilite oranlarına sahip dokuz Holştayn boğadan alınan ve dondurulan spermalar in vitro fertilizasyon amacıyla kullanılmıştır. İn vitro fertilizasyon için Brackett ve Oliphant mediumu ile Direkt yıkama metodu ve 5-6 saat inkubasyon periyodu; in vitro embriyo kültürü için de Charles Rosenkrans mediumu kullanılmıştır. Kültür periyotlarında %95’in üzerinde bağıl nem, 39°C, %5 CO2 içeren bir kültür ortamı sağlanmıştır. Toplam 2519 A ve B kalite oosit in vitro embriyo elde etme sürecine alınmıştır. Sonuç olarak yarıklanma ve blastosiste ulaşma oranları bakımından boğalar arasında önemli düzeyde istatistiki farklılık bulunmuştur (P0.05). Bu sonuçlar boğaların değişik düzeyde in vitro fertilizasyon ve embriyo üretim yeteneklerine sahip olduklarını ve in vitro embriyo üretim başarısı için erkek faktörünün belirleyici olduğunu göstermektedir.

In vitro embriyo üretimine boğa etkisinin araştırılması

The aim of the study was to show whether there were some differences among 9 Holstein bulls and within their replications on their ability of in vitro fertilization for in vitro embryo production as cleavage and coming into the blastocyst stage. Semen collected and frozen from nine Holstein bulls with satisfactory in vivo fertilization capabilities for artificial insemination was used for in vitro fertilization. Direct washing method by Brackett and Oliphant medium and 5 or 6 h incubation period were used for in vitro fertilization. Charles Rosenkrans medium was used for in vitro embryo culture. An atmosphere with a higher than 95% relative humidity, 39°C, 5% CO2 and was used for all in vitro embryo production processes. Totally 2519 A and B quality oocytes were treated for in vitro embryo production. As a result statistically significant (P<0.05) variation was found for cleavage and blastocyst development among bulls. However there was no significant difference (P>0.05) between the replications for each bull as cleavage and coming into blastocyst stage. The results showed varied capabilities of bulls for in vitro fertilization and embryo production and male factor can affect success of in vitro embryo production.

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1. Gordon I: Laboratory Production of Cattle Embryos. CAB International Ltd. Co., University Press, Cambridge, UK, 1994.
2. Eyestone WH, First NL: Variation in bovine embryo development in vitro due to bulls. Theriogenology, 31 (1): 191, 1989.
3. Hillery FL, Parrish JJ, First NL: Bull specific effect on fertilization and embryo development in vitro. Theriogenology, 33 (1): 249, 1990.
4. Shi DS, Lu KH, Gordon I: Effects of bulls on fertilization of bovine oocytes and their subsequent development in vitro. Theriogenology, 33 (1): 324, 1990.
5. Graham JK, Moce E: Fertility evaluation of frozen/thawed semen. Theriogenology, 64 (3): 492-504, 2005.
6. Schneider CS, Ellington JE, Wright Jr RW: Relationship between bull field fertility and in vitro embryo production using sperm preparation methods with and without somatic cell co-culture. Theriogenology, 51 (6): 1085-1098, 1999.
7. Moce E, Graham JK: In vitro evaluation of sperm quality. Anim Reprod Sci, 105 (1): 104-118, 2008.
8. First NL, Parrish JJ: In-vitro fertilization of ruminants. J Reprod Fertil Suppl, 34, 151- 165, 1987.
9. Kanagawa H, Shimohira I, Saitoh N: Manual of Bovine Embryo Transfer National Livestock Breeding Center MAFF, JICA, Japan, 1995.
10. Rosenkrans CF, First NL: Effect of free aminoacids and vitamins on cleavage and developmental rate of bovine zygotes in vitro. J Anim Sci, 72 (2): 434-437, 1994.
11. Niwa K, Ohgoda O: Synergistic effect of caffeine and heparin on in- vitro fertilization of cattle oocytes matured in culture. Theriogenology, 30 (4): 733-741, 1988.
12. Brackett BG, Zuelke KA: Analysis of factors involved in the in vitro production of bovine embryos. Theriogenology, 39 (1): 43-64, 1993.
13. Galli C, Lazzari G: Practical aspects of IVM/IVF in cattle. Anim Reprod Sci, 42 (1): 371-379, 1996.
14. Lu KH, Gordon I, Gallagher M, McGovern H: Pregnancy established in cattle by transfer of embryos derived from in vitro fertilization of follicular oocytes matured in vitro. Vet Rec, 121 (11): 259-260, 1987.
15. Al Naib A, Hanrahan JP, Lonergan P, Fair S: In vitro assessment of sperm from bulls of high and low fertility. Theriogenology, 76 (1): 161- 167, 2011.
16. Otoi, T, Tachikawa S, Kondo S, Suzuki T: Effects of different lots of semen from the same bull on in vitro development of bovine oocytes fertilized in vitro. Theriogenology, 39 (3): 713-718, 1993.
17. Leibfried-Rutledge ML: Factors determining competence of in vitro produced cattle embryos. Theriogenology, 51 (2): 473-485, 1999.
18. Kato H, Iritani A: In vitro fertilization in cattle. Mol Reprod Develop, 36 (2): 229-231, 1993.
19. Takahashi Y, Hishinuma M, Matsui M, Tanaka H, Kanagawa H: Development of in vitro matured/fertilized bovine embryos in a chemically defined medium: Influence of oxygen concentration in the gas atmosphere. J Vet Med Sci, 58 (9): 897-902, 1996.
20. Leibfried-Rutledge ML, Critser ES, Eyestone WH, Northey DL, First NL: Developmental potential of bovine oocytes matured in vitro or in vivo. Biol Reprod, 36 (2): 376-383, 1987.
21. Wiemer KE, Watson AJ, Polanski V, Mc Kenna AI, Fick GH, Schultz GA: Effects of maturation and co-culture treatments on the developmental capacity of early bovine embryos. Mol Reprod Develop, 30 (4): 330-338, 1991.
22. Tammassia M, Heyman Y, Lavergne Y, Richard C, Gelin V, Renard JP, Chastant-Maillard S: Evidence of oocyte donor cow effect over oocyte production and embryo development in vitro. Reproduction, 126 (5): 629-637, 2003.
23. Palma GA, Sinowatz F: Male and female effects on the in vitro production of bovine embryos. Anat Histol Embryol, 33 (5): 257-262, 2004.