• Korean Journal of Veterinary Research
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• v.28 no.1
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• pp.203-211
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• 1988

The present study was carried out to investigate the estrous behavior and vaginal smear after induction of estrus with exogenous hormones in the premature, metestrous and anestrous bitches. A total of 21 bitches(Mixed breed: 16, Jindo breed: 5), from 10 months to 5 years of age and weighing 8 to 15 kg was studied the change of vaginal smear and the estrous behavior before and after induction of estrus. The results obtained are as follows: 1. In the treatment A(They were given the $PGF_2{\alpha}$, estrone, estradiol-$17{\beta}$, PMSG and HCG) proestrus commenced in $10.16{\pm}1.44$($Mean{\pm}SEM$) days after treatment, The mean duration of proestrus, and estrus was $7.50{\pm}1.44$ and $13.50{\pm}3.44$ days, respectively. In the treatment B(They were given the PMSG and HCG) proestrus commenced in $5.53{\pm}0.59$ days after treatment. The mean duration of proestrus and estrus was $6.60{\pm}0.71$ and $14.60{\pm}1.14$ days, respectively. 2. All of the 6 bitches in the treatment A showed vulval swelling and vaginal discharge, 14 of the 15 bitches in the treatment B showed vulval swelling and vaginal discharge. However, all of the treatment A and B showed male acceptance. 3. The main change of vaginal smear in proestrus and estrus after induction of estrus was a increase in the proportion of anuclear and superficial cells associated with a decrease in small intermediate and parabasal cells. By the estrous behavior and vaginal smear the estrus was induced in all the premature, metestrous and anestrous bitches.

• Korean Journal of Animal Reproduction
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• v.14 no.1
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• pp.23-30
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• 1990

These studies were carried out to investigate the effects of fetal calf serum(FCS), estrous porcine serum(EPS), porcine follicular fluid(PFF), hormone and matured cumulus cell(MCC) on in vitro maturation and fertilization of porcine follicular oocytes. The ovaries and testes were obtianed from slaughtered Landrace sow and boars, respectively. The follicular oocytes surrounded with cumulus cells were recovered by aspirating follicular fluid from the visible follicles of diameter 3~5 mm and the semen were prepared from boar's epididymal cauda. The follicular oocytes were cultured in TCM-199 medium containing hormones, FCS, EPS, PFF and MCC for 48hrs. in a incubator with 5% CO2 in air at 36$^{\circ}C$ and then matured oocytes were again cultured for 18~20 hrs. with $1.5\times$106/ml motile capacitated sperm in the modified Tyroide solution containing 100$\mu\textrm{g}$/ml of heparin. The results obtained in these experiments were summarized as follows : 1. The maturation and fertilization rate of the follicular oocytes, cultured in the TCM-199 medium supplemented with 10% FCS and PMSG＋HCG were 55.6~64.5% and 33.3~37.1%, respectively. 2. The maturation and fertilization rate of the follicular oocytes cultured in the TCM-199 medium supplemented with 20% EPS and PMSG＋HCG were 50.0~55.0% and 30.3~33.3%, respectively. 3. The maturation rate(59.0~64.2%) and fertilization rate(34.8~39.3%) of follicular oocytes cultured in TCM-199 medium supplemented 20% FCS and 50% PFF were higher than those of follicular oocytes cultured in TCM-199 medium supplemented with 5%, 10% and 15% FCS and 10% and 50% PFF. 4. The maturation rate(60.0%) and fertilization rate(40.0%) of follicular oocytes cultured in TCM-199 medium supplemented with 20% FCS and granulosa cell (1$\times$106/ml) were significantly higher than those of fiollicular oocytes cultured in TCM-199 medium supplemented with 5%, 10% and 15% FCS and granulosa cell.

• Journal of Aquaculture
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• v.9 no.3
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• pp.287-291
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• 1996

[ $F_2$ ] generation of gynogenetic diploids were produced in olive flounder, Paralichthys olivaceus, by brother-sister mating with full-sibling gynogenetic diploids ($F_1$). The induced ovulations and spawnings were conducted by using intraperitoneal injections of HCG (2,000 IU/kg body weight) with photoperiod controls. Floating rates of artificially ovulated eggs were ranged from 22.9 to $65.7\%$. Fertilization and hatching rates were ranged from 69.0 to $86.2\%$ and 36.8 to $85.8\%$, respectively. All of those three rates between two experimental groups were not significantly different (P>0.05). Average survival rates of $F_2$ generation of 40-day-old 효nogenetic diploid larvae were slightly lower than those of controls (P<0.05), however growth rates were much higher than those of their diploid controls.

• Korean Journal of Animal Reproduction
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• v.16 no.4
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• pp.301-310
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• 1993

This experiment was performed to develop simple practical methods for short term preservation of rabbit embryos. A total of 55 cross bred does were superovulated by intramuscular injection of PMSG and HCG. Embryos were recovered at 25~30 hrs, 60~65 hrs and 80~85 hrs after mating and selected by morphological examination. Four cell stage, morulae and blastocyst embryos were stored in PBS enrich with 1, 10, 20 and 40% heat-treated FCS at 4, 20, 30 and 37$^{\circ}C$, respectively. Embryos were examined morphologically at 24, 48 and 72 hrs following storage. The result obtained in this experiment were summarized as follows: The superovulation was induced by PMSG 200 IU and HCG 100 IU. The average number of ovulation points and embryos recovered by collection time were 19.0, 15.6(25~30 hr), 17.3, 13.5(60~65 hr) and 19.2, 14.4(80~85 hr), respectively. And recovery rates of embryos recovered at 25~30 hr, 60~65 hr and 80~85 hr after mating were 62.8%(4 cell), 84.7%(morulae) and 79.6%(blastocyst), respectively. On the other hand, the average number of ovulation points collected by the no, of operations for the repeated collection was 17.3(60~65 hr), 19.2(80~85 hr) in 1st and 9.4(60~65 hr), 10.6(80~85 hr) in 2nd surgery, respectively. There was a significant decrease(P<0.05) in the number of ovulation points the 2nd surgery as compared to the 1st surgery. All of the 4-cell stage embryos stored at 4$^{\circ}C$ for 48 hrs showed the same morphology throught the storage period, on the contrary, 4-cell stage embryos stored at 2$0^{\circ}C$ and 3$0^{\circ}C$ for 48 hrs showed degeneration embryos and stored at 37$^{\circ}C$ for 24 hrs showed degeneration embryos. Morulae and blastodcyst stored at 4, 20, 30 and 37$^{\circ}C$ for 24 hrs showed degeneration embryos. All of the blastocyst stored at 37$^{\circ}C$ for 72 hrs showed degeneration embryos.

• Korean Journal of Veterinary Research
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• v.29 no.3
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• pp.253-262
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• 1989

This study was performed to investigate the patterns of progesterone secretion after induction of estrus in premature, metestrous and anestrous bitches. A total of 22 bitches were used. Of them 18 bitches were treated with hormone to induced estrus and 4 bitches were untreated and served as controls. Estrus was induced with $PGF_{2{\alpha}}$, estrone, estradiol-$17{\beta}$, PMSG and HCG(Treatment A), and with PMSG and HCG(Treatment B). Blood samples were collected via the cephalic vein at 2 to 5 days interval. Blood samples were centrifuged (1,200g, 10min.) within 30 minutes after collection and plasma was stored at $-20^{\circ}C$ until analyzed for the progesterone concentrations. Plamsa progesterone concentrations were measured by radioimmunoassay. The results of estrous induction were determined by estrous signs, ovarian response, egg recovery and progesterone patterns. The results obtained were as follows; 1. All bitches in treatment A showed estrous signs, however the ovarian response and egg recovery were not detectable and the levels of progesterone were nearly same as before. 2. In the treatment B, premature and metestrous bitches showed only estrous signs, however 5 of 7 anestrous bitches (71.4%) showed estrous signs, ovarian response and changes of progesterone levels. In conclusion, clinical estrous behavior can be induced during any phase of the estrous cycle, but ovulation should be induced only if induction occur approximately 4 months or more after the previous estrus.

• Korean Journal of Animal Reproduction
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• v.12 no.3
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• pp.132-140
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• 1988

These experiments were carried out to examine the development capacity of mouse blastomers separated from 2 to 8-cell stage mouse embryos. The female ICR and C3H mice were subjected to supervolution by intraperitoneal injection of PMSG and HCG and then mated with males of the same strain. Embryos were flushed from oviducts and uteri on a proper time after injection of HCG. After removal of zona pellucida with 0.5% pronase, each embryos were separated into 1/2, 1/4, 2/4, 1/8, 2/8 and 4/8 embryos by pipetting or a fine glass needle in Ca2＋$.$Mg-2＋ free Hoppe& Pitts medium containing 0.02% EDTA. Splitted embryos were cultured in Hoppe & Pitts medium for 48h to 72h. The embryos developed to blastocyst were transferred to recipients on 2 or 3 days of pseudopregnancy. On the other hand, a monozygotic pairs of 1/2 embryos developed to blastocyst after 48h in vitro culture were transferred to recipients on 2 days of pseudopregnancy or pregnancy. The results obtained were summarized as follows. 1. Success rates of separation of blastomeres from 2-, 4- and 8-cell embryos were 91.7%, 68.5-92.4% and 60.8-90.6%, respectively. 2. Development rates of various type of blastomeres to blastocyst after 72h in vitro culture were ranged 64.7-87.1%. 3. Blastocysts obtained after 48h in vitro culture were transferred to recipients on 2 or 3 days of pseudopregnancy. The production rates of live fetuses after transfer on 2 days, only 1/2, 2/4 and 4/8 embryos, were 13.2%, 13.5% and 17.2%, respectively and those of embryos transferred on 3 days were 11.8%, 9.6% and 11.5%, respectively. However, the production rates of live fetuses 1/2 embryos following 72h in vitro culture and transfer to recipients on 2 or 3 days of pseudopregnancy were 7.7% and 12.5%, respectively. 4. From 29 and 31 pairs of 1/2 embryos transferred to recipients on 2 days of pseudopregnancy or pregnancy, 4 sets of monozygotic twins were produced from only pregnant recipients.

• Journal of Aquaculture
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• v.3 no.1
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• pp.25-30
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• 1990

Adult channel catfish, Ictalurus punctatus, mature but do not ovulate and spawn in ponds in Korea. Ovulation and natural spawning were induced in the breeding season by increasing water temperature or injecting one to three doses of both 4.4 mg dried carp pituitary or 1,100 IU human chorionic gonadotropin(HCG) per kg body weight at 24 hours intervals when plural doses were employed. The increasing higher temperature (from $24^{\circ}C$ to $30^{\circ}C$) resulted in 4 times greater ovulation than lower increase of temperature (from $24^{\circ}C$ to $27^{\circ}C$). In overall performance, dried carp pituitary gave $78.6\%$ ovulation and HCG induced $66.6\%$ ovulation. None of the control fish spawned in any of the experiments.

• Journal of Microbiology and Biotechnology
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• v.9 no.5
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• pp.541-547
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• 1999

Constructions of a transfer vector and a recombinant baculovirus using the thymidine kinase gene of the Herpes simplex virus type 1 strain F (HSV -1) were carried out. Newly cloned transfer vector, pHcgXIIIB, was constructed by insertion of the glycoprotein gX gene signal peptide sequence of Pseudorabies virus into the baculovirus vector pHcEV-IV. The gX sequence was inserted just downstream from the promoter for the polyhedrin gene of the Hyphantria cunea nuclear polyhedrosis virus (HcNPV). HSV-1 thymidine kinase(tk) gene (1.131 kb) was used as a candidate gene for transferring into the baculovirus expression system. The tk gene was inserted into a BamHI site downstream from the gX sequence-promoter for the polyhedrin gene in the pHcgXIIIB transfer vector and was transferred into the infectious lacZ-HcNPV expression vector. Recombinant virus was isolated and was named gX-TK-HcNPV. The recombinant virus produced a 45 kDa gX-TK fusion protein in Spodoptera frugiperda cells, which was confirmed by Western blot analysis. Microscopic examination of gX-TK-HcNPV-infected cells revealed normal multiplication. Fluorescent antibody staining indicated that the gX-TK fusion protein was present in the cytoplasm. These results indicated that the transfer vector successfully transferred the gX-tk gene into the baculovirus expression system.

• Korean Journal of Animal Reproduction
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• v.17 no.1
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• pp.57-68
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• 1993

This stduy was carried out to find a reliable method for the production of in vitro fertilized embryos having more excellent development capacity and freezability in the rabbit and cattle. The greatest number of rabbit oocytes was recovered 6hrs after HCG injection(P<0.05). The maturation rate in vitro was slightly higher in the oocytes(6-h-oocytes) from 6h than those (8-h-oocytes)from 8 hrs after HCG injection and the beneficial effect of FSH during oocyte maturation was significantly great in the oocytes from large follicles. The cleavage rate into 2-to-6-cell stage was not differ between the 6-h-oocytes and 8h-oocytes, but the cleavage of these oocytes was greatly promoted by FSH addition to maturation medium and the cleavge of 8-h-oocytes matured without FSH was significantly low. The embryo development into 16-cell to morula was not promoted by the co-culture with rabbit oviduct epithelial cells. The freezability by embryo stages was ovidusly high at 4-cell and morula stage in 6-h-oocytes and the viability of 16-cell embryos from 8h-oocytes was similar to that of morula stage. The implantation sites after surgical tranfer of fresh rabbit embryos were not implanted. In bovine experiment, the in vitro development into 16-cell and morula after in vitro maturation and fertilization in the follicular oocytes was slightly improved by the co-culture with granulosa cells compared to that with oviduct epithelial cells and the frozen-thawed viability rate of these embryos ranged from 14 to 40%. The excellent fresh embryos were transferred nonsurgically to 6 recipients, but were not pregnant.

• Korean Journal of Veterinary Research
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• v.32 no.4
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• pp.663-669
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• 1992

This experiment was carried out to find out the best condition for the parthenogenetic activation of mouse eggs by treating ethanol and hyaluronidase. For the parthenogenetic activation of eggs with ethanol, cumulus cell enclosed or denuded eggs were treated with 7% ethanol in D-PBS for 5, 7 or 9 minutes. For the activation of eggs with hyaluronidase, the eggs with cumulus masses were released into D-PBS with 100 unit hyaluronidase and treated for 10, 12 or 13 minutes. All of the treated eggs were incubated in BMOC-3 solution for 5 hours at $37^{\circ}C$ at an atmosphere of 5% $CO_2$ in air. The types of parthenogenetic eggs were morphologically classified into haploid, diploid, immediate cleavage eggs under an inverted microscope. The results obtained in this experiment were summarized as follows ; 1. High activation rate(99%) had been achieved by treating the eggs with 7% ethanol for 7 minutes. 2. With 100 IU hyaluronidase, high activation rate (94%) had been achieved by treating for 12 minutes. 3. The most frequent type of parthenogenetic eggs activated with ethanol or hyaluronidase was haploid (p<0.05). 4. The eggs collected from 18 to 22 hours post HCG injection showed higher activation rate than the eggs collected at 16 hours post HCG injection. 5. No significant difference (p>0.05) in activation rate was shown in strain of mouse and in presence of cumulus cells.