Culture of preantral follicles has important biotechnological implications through its potential to produce large quantities of oocytes for embryo production and transfer. The objective of this study was to determine the comparison of different isolation method of mouse preantral follicles, and to examine in vitro development of mouse preantral follicles isolated by different method. Preantral follicles were mechanically or enzymatically extracted from mouse ovaries. Mechanical isolation method used fine gauge needles and enzymatic method of isolating follicles used 1 mg/ml collagenage (Type IA) and 0.2 mg/ml DNase Ⅰ in Leibovitz L-15 medium. The solution containing Leibovitz L-15 medium, enzyme and ovary fragments was incubated at 37℃ for 30 min. The selection criteria are as follows: primary follicle of 75 to 99 μm, early secondary follicle of 100 to 125 μm and late secondary follicle of 126 to 150 μm in diameter. The recovered preantral follicles were cultured for 10 days in alpha-minimal essential medium (α-MEM) + 5% FBS + ITS + 100 mIU/ ml FSH. The collected primary follicles by enzymatic treatment were higher than mechanical method. Others stage preantral follicle by mechanical isolation were higher than enzymatic method. After 10 days of culture, no statistical differences were shown in survival rates of preantral follicle among the 2 culture groups. The metaphase Ⅱ rates of the oocytes were significantly higher (p<0.05) in mechanical method (17.8%) than in enzymatic method (5.1%). These results suggest that the isolation method of choice depends on the target stage preantral follicles and mechanical isolation is an optimal method of preantral follicles in a culture of mouse preantral follicle.

The objective of this study was to compare the accuracy analysis and the effect of field application of a newly developed automatic heat detector in dairy cows. From 2009 to 2010, we used 48 Holstein cows (mounting cows : 38 heads, standing cows : 10 heads) raised in experimental barn of National Livestock Research Institute (RDA) for the accuracy analysis of automatic heat detector, and 14 Holstein cows raised in three commercial dairy farms of Cheonan and Pochun area for comparison of the effect of field application. The accuracy of response in cows attached with automatic heat detector was 86.8% (33/38) displayed on board when mounting activity observed, and 100% (10/10) when standing activity observed, and on average, 90.0% (43/48) displayed on board. The accuracy of automatic heat detector in on-farm test was 85.7% (12/14), and conception rate was 75.0% (9/12).

The aim of this study was to evaluate the effect of α-tocopherol on blastocysts development and subsequent cryosurvival of the vitrification. The α-tocopherol(0, 100, 200, 400 μM) was added in to culture medium for the bovine embryos. The blasocysts from the α-tocopherol and untreated control groups were then frozen-thawed, and their cryosurvival was assessed by in vitro culture for 48 h. There were no differences in the overall cleavage rate(56.14±4.66, 58.18±4.70, 62.97±6.86 and 51.17±7.28) among four treatment groups. However, in blastocyst development and total cell number were significantly higher in α-tocopherol 200 μM(38.60±7.12; 106.33±3.50) to culture medium than other treatment groups(29.30±5.24, 31.60±7.12 and 26.37±4.18; 101.36±5.12, 97.27±2.87, and 91.23±7.52 respectively). Before and after vitrification, the total cell number and blastocyst development of embryo were significantly higher in July to August than September to October. In conclusion, addition of α-tocopherol 200 μM to in vitro bovine embryo culture medium was beneficial for improving embryo quality by decreasing the embryo damage blsstocysts cell number and improving the tolerance of the embryos to cryopreservation.

The objective of this study was to investigate the relationship between estrous expression, body condition score (BCS), blood urea nitrogen (BUN) and number of transferable embryos for the purpose of improving reproductive performance in blood of Hanwoo donors. Sixty, at random stages of the estrous cycle, received a CIDR. Four days later, the animals were superovulated with a total of 28AU FSH (Antorin, 2AU=1 ml) administered twice daily in constant doses over 4 days. On the 3th administration of FSH, CIDR was withdrawn and 25 mg PGF2α was administered. Cows were artificially inseminated twice after estrous detection at 12 hr intervals. The cows received 100 μg GnRH at the time of 1nd insemination. Embryos were recovered 7 or 8 days after the 1st insemination. The estrous inducement rate and estrous expression rate were significantly lower for cows with BCS below 2.25 than for cows with BCS above 2.25. There was 50.0% of rate of mounting in cows with BCS below 2.25 whereas the rate of mounting was markedly increased in cows with BCS above 2.25 (94.1% and 89.5% for BCS 2.25～2.75 and BCS above 2.75 cows, respectively). Cows with BCS ＜2.25, 2.25～2.75 and ≥2.75 had number of transferable embryos of 4.5±0.7, 5.9±1.8 and 5.6±2.3 respectively.

The objective of this study was to investigate the relationship between concentration of urea nitrogen, glucose, cholesterol and number of transferable embryos for the purpose of improving reproductive performance in blood of Hanwoo donors. Fifty five, at random stages of the estrous cycle, received a CIDR. Four days later, the animals were superovulated with a total of 28AU FSH (Antorin, 2AU=1 ml) administered twice daily in constant doses over 4 days. On the 3th administration of FSH, CIDR was withdrawn and 25 mg PGF2α was administered. Cows were artificially inseminated twice after estrous detection at 12 hr intervals. The cows received 100μg GnRH at the time of 1nd insemination. Embryos were recovered 7 or 8 days after the 1st insemination. Cows with BUN ＜10, 11～18 and ≥19 mg/dl had number of transferable embryos of 4.3±1.3, 5.8±1.8 and 4.7±2.1 respectively. The mean numbers of total ova from < 10 and 10≤ of corpora lutea(CL) was 8.9 and 14.3, respectively. The number of transferable embryos differed between < 10 and 10≤ CL was 4.8 and 5.6, respectively.

This study was conducted to investigate the survival rate of frozen-thawed spermatozoa in equine by glycerol concentration and freezing speed. Two stallions (1 Thoroughbred-13 year old and 1 Arab-7 year old) bred in Korea Racing authority was examined for 1 times in a couple of weeks. Semen was collected by condom method standing heated mare and were centrifuged 650 g for 15 min. and isolated the seminal plasma. Thick fraction of semen was diluted EDTA-Lactose-egg yolk diluents to 1:1 and contained in 0.5 ml straw as 6～14×107cells/ml. Final concentrations of glycerol were 3, 5 and 7% in cryopreseved diluents and added 4 times for 2 hours equilibration. For the freezing, equilibrated straws were located 3 or 5 cm above LN2 gas for 5 or 10 min. Survival rates of pre-frozen sperm were 65.0±13.2%, 68.3±10.4%, 66.7±11.5% and post-frozen were 53.3±23.1%, 45.0±15.0%, 50.0±18.0% in 3, 5, 7% glycerol concentration, respectively. There was no difference between glycerol concentrations. Survival rates of frozen-thawed sperm on freezing speed were 36.7±10.4%, 40.0±7.1%, 30.0±13.2% at 3 cm－5 min and 33.3±11.5%, 31.7± 2.9%, 21.7±10.4% at 3 cm－10 min in 3, 5, 7% glycerol concentration, respectively. Survival rates of frozen-thawed sperm on freezing speed were 43.3±15.3%, 32.0±17.9%, 22.3±15.7% at 5cm－5 min and were 47.5±15.0%, 43.3±12.6%, 48.3±15.3% at 5cm－10 min in 3, 5, 7% glycerol concentration, respectively. There were significantly different between groups (p<0.05). These results suggest that glycerol concentration did not affect cryopreservation of stallion semen within 3～7% but freezing speed affects. In our experiment, the best cryopreservation condition was at 5 cm above LN2 gas for 10 min for pre-freezing and 7% of glycerol concentration. These results lead to commercial AI with frozen-thawed stallion semen.

This study was performed to investigate the characteristics within ages and freezing tolerance of spermatozoa in Jindo Dog. Experimental animals were selected 12 herds within 1～8 year’s old and collected semen for 2 times in a week. Collected semen was evaluated whole volume and sperm number with CASA system (SIAS, Medical Supply, Korea). Then seminal plasma were separated and diluted with modified Tris-egg yolk extender and added 4, 6 and 8% glycerol for 4 times to final concentration and equilibrated for 1.5 hrs. Before and after freezing, equilibrated semen were evaluated the survival rates. Total volume of sperm at 1～2 year old group is as 5.2×108 cells/ ml largest and there were no significance among groups. The motility of 1～2 year old group is highest as 90.9% and there were significance among groups. Abnormal sperm showed similar among groups. The survival rate in terms of pre-freezing and post-freezing were decreased all levels of glycerol and reveled 87.0% to 64.5% in 4%, 87.5% to 51.9% in 6% and 73.4% to 29.7% in 8%, there were significant difference among the groups (p<0.05). These results suggest that the optimal sperm-freezing methods in Jindo Dog are utilized with modified Tris egg-yolk extender with 4% glycerol and were improve the reproductive activity by these methods.

The objective of this study was investigate the superovulation treatment and to relate concentrations of blood urea nitrogen(BUN) in Hanwoo donors. Thirty six, at random stages of the estrous cycle, received a CIDR. Four days later, the animals were superovulated with a total of 28AU FSH (Antorin, 2AU=1 ml) administered twice daily in constant doses over 4 days. On the 3th administration of FSH, CIDR was withdrawn and 25 mg PGF2α was administered. Cows were artificially inseminated twice after estrous detection at 12 hr intervals. The cows received 100 μg GnRH at the time of 1st insemination. Embryos were recovered 7 or 8 days after the 1st insemination. Cows with BUN ＜10, 11～18 and ≥19 mg/dl had return of estrus of 34.6, 30.5 and 30.4 days respectively. Return of estrus after superovulation treatment was not significantly lower for cows with blood urea nitrogen (BUN) above 10 mg/dl than for cows with BUN below 10 mg/dl. Cows with BUN ＜10, 11～18 and ≥19 mg/dl had number of transferable embryos of 3.2±1.2, 5.4±1.9 and 4.1±2.1 respectively.