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KMID : 0900920020260010001
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Korean journal of Animal Reproduction
2002 Volume.26 No. 1 p.1 ~ p.7
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Effect on InVitro Fertilization of Pig Oocytes Matured in Different InVitro Maturation Media according to Sperm Concentration of Liquid Boar semen
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Park Chang-Sik
Lee Young-Joo
Ko Hyeon-Jin
Seo Kil-Woong
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Abstract
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The present study was carried out to investigate the effects of maturation media on penetrability of pig oocytes by liquid boar sperm coincubated with different sperm concentrations in a modified Trisbuffered medium (mTBM). Follicular oocytes collected from ovaries of prepubertal gilts were matured in a modified TCM-199 (mTCM-199) medium, modified Waymouth MB 752/1 (mWaymouth MB 752/1) medium or NCSU-23 medium. Oocytes (30~40) were transferred into each well of a Nunc 4-well multidish containing 0.5 mell maturation medium. The spermich portion of ejaculates with greater than 90% motile sperm were used in the experiment. The semen was cooled 22 to 24^{circ}C over 2 h period. The semen was diluted with Beltsville Thawing Solution (BTS) extender at room temperature to give 2times10^{8} sperm/mell in 100 mell plastic bottle. Liquid boar semen of 30 mell in 100 mell plastic bottle was kept at 17^{circ}C far 5 days. The sperm with greater than 70% motility after day 5 of storage were used for in-vitro fertilization (IVF). After 44 h maturation of immature oocytes in 5% CO_2in air at 38.5~{circ}C, cumulus cells were removed and oocytes (30~40) were coincubated for 6 h in 0.5 mell mTBM fertilization medium with five different (1times10^{6} , 2times10^{6} , 4times10^{6} , 6times10^{6}, 10times10^{6} mell) sperm concentrations. At 6 h after IVF, oocytes were transferred into 0.5 mell NCSU-23 culture medium fur further culture of 6 h. At 12 h after IVF, sperm penetration, polyspermy and male pronuclear formation of oocytes were evaluated. Oocytes of NCSU-23 maturation medium decreased polyspermy and increased male pronuclear formation compared to those of mTCM199 and mWaymouth MB 752/1 maturation media. Of oocytes matured in NCSU-23 medium and inseminated in mTBM medium with 2~4times10^{6} mell sperm concentrations, 50.8~50.9% showed sperm penetration, 13.3~19.5% polyspermy and 43.9~45.4% male pronuclear formation. In conclusion, we found out that oocytes matured in NCSU23 medium and inseminated in mTBM medium showed superior invitro fertilization compared to those matured in mTCM199 and mWaymouth MB 752/1 maturation media and inseminated in mTBM medium. The optimum sperm concentrations for in-vitro fertilization of oocytes matured in NCSU-23 medium by liquid boar semen stored at 17^{circ}C for 5 days were 2~4times10^{6} mell.
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KEYWORD
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In-vitro fertilization, Pig oocytes, Maturation medium, Liquid boar semen, Male pronuclear formation
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