Transgenic chickens have been spotlighted as an highly potent bioreactor for their fecundity, short generation time, and eggs associated with mass production of protein. In this study, we generated transgenic chickens exhibiting oviduct specific expression of human growth hormone fused to human transferrin for oral administration. Gene of the modified growth hormone located at downstream ovalbumin promoter (∼3.6 kb) was introduced to stage X blastodermal cell employing retrovirus vector system. Several transgenic chickens were successfully generated. However, genomic analyses showed unexpected deletion within the transgene. The modification of the transgene seemed to occur during germ cell formation because the deletion was detected only from the sperm DNA of the G0 founder animal. There was no evidence of deletion in the somatic cell DNA samples of the same chicken. Consequently, same pattern of the deletion was confirmed in both somatic and germ cells of the G1 progeny.
The objective of this study was to develop of semen transport system for cryopreservation and fertility in bull sperm. The ejaculated semen were diluted with Triladyl containing 20% egg-yolk for transportation. Diluted semen was transported by three methods that there were wrapping tissue (Tissue), sinking under 30℃ water (Water) and sinking between warm water and air (Air) methods. Semen was transported within 2 hours in 0.3℃. For this study, the freezing of diluted semen were added with Triladyl containing 20% egg-yolk. And frozen-thawed sperm were estimated with SYBR14/PI double stain for viability, FITC-PNA/PI double stain for acrosome reaction analysis and Rhodamine123 double stain for mitochondrial intact assessment. In results, live sperm (SYBR+/PI-) in Air treatment group (43.3±4.7%) was significantly (p<0.05) higher than other treatment groups (Tissue: 16.3±2.7% and Water: 27.5± 3.1%), dying sperm (SYBR+/PI+) in Air treatment group (55.6±4.7%) was significantly lower than other treatment groups (Tissue: 77.6±3.2% and Water: 67.6±3.3%) (p<0.05). Acrosome reaction in Air treatment group (0.2±0.1%) within live sperm (PI negative region) was significantly (p<0.05) lower than other treatment groups (Tissue: 0.7±0.2% and Water: 0.5±0.1%), the acrosome reaction in Air treatment group (28.6±2.8%) within all sperm also was significantly lower than other treatment groups (Tissue: 44.2±1.8% and Water: 36.2±2.0%) (p<0.05). And mitochondrial intact in Air treatment group within live (97.1±0.4%) and all (61.9±3.3%) sperm were significantly higher than other treatment groups (Tissue: 85.2±3.3%, Water: 87.8±2.9% within live sperm and Tissue: 49.28±3.7%, Water: 42.0±3.1% within all sperm) (p<0.05). Therefore, we suggest that transportation by sinking method between warm water and air was beneficial to improvement of fertility in frozen-thawed in bull semen.
An uterus is female reproductive tract organ that affected estrus cycle. During a various changes occur at uterus in estrus cycle, one of them is body fluids secretion be called uterine fluid. Therefore, the objective of this study was to investigate the changes of protein patterns using two-dimensional gel electrophoresis in uterus fluids during the follicular and luteal phases in estrus cycle of pigs. In changes of protein spots were confirmed during the follicular and luteal phases. The 136 spots were expressed in follicular phase, the 57 spots of them showed reproducibility. On the other hand, the 140 spots were expressed in luteal phase, the 73 spots of them showed reproducibility. Also, spots expressed in follicular phase were number 69 and 94 spots and spots expressed in luteal phase only were number 156, 157, 184~187, 190 and 191 spots. The spots which of higher expression levels in the luteal phase than in follicular phase were number 76 and 79 spots. In conclusion, the spots expressed in follicular and luteal phases were confirmed with difference levels and these differences are function of RNA resolving, protein synthesis and cytoskeletal architecture.
Until recently the most popular tetracycline-inducible gene expression system has been the one developed by Gossen and Bujard. In this study, we tested the latest version of same system and the results are summarized as follows: Compared with previous one, the difference of new system are minor changes of nucleotide sequences in transactivator and tetracycline response element (TRE) regions. Sensitivity to the doxycycline (a tetracycline derivative) was improved. Leakiness of GFP marker gene expression in non-inducible condition was significantly decreased. Higher expression of the marker gene was observed when the cells were fed with doxycycline- containing medium. Optimal insertion site of woodchuck posttranscriptional regulatory element (WPRE) sequence which was known to increase gene expression was different depending on the origin of cells. In chicken embryonic fibroblast, location of WPRE sequence at 3’ end of TRE resulted in the highest GFP expression. In bovine embryonic fibroblasts, 3’ end of transactivator was the best site for the GFP expression.
The aim of this study was to investigate the effects of dietary supplementation of pelleted-Italian Ryegrass (IRG) as a source of fiber on reproduction performance in pregnant sows. A total of 24 pregnant sows were randomly assigned to four dietary treatments, which was given a corn-soybean diet with 0%, 10%, 20%, and 30% pelleted-IRG from 105 days prepartum to 7 days postpartum. During experimental period, the sows fed the IRG supplemented diet showed the lower feed intake than the sows fed the control diet (p<0.05). The changes of body weight in sows from initial to pre- and/or post-partum was significantly smaller in sows fed the IRG supplemented diet than control group. It is thought that the lower weight gain in IRG supplemented groups is caused by low feed intake. Although there was no significant difference, sows fed the IRG supplemented diet tended to increase the litter size and birth weight in piglets compared with sows fed the control diet. This result suggests that the dietary supplementation of IRG has the positive effects to improve the reproductive performance in sows. But, the excessive feeding of IRG to sows might cause to retard the days of return to estrus, and decrease the contents of solid, milk protein, and milk fat in colostrum. Thus, the addition of about 10% IRG is desirable to increase the reproductive performance. Meanwhile, the feeding energy diet is better effective than feeding the fiber diets to improve overall productivity in sows after postpartum.
The objective of this study was to compare the effects of the levels of inbreeding on body weight traits between two breed populations, Hanwoo and Korea Brindle cattle. Birth weight (BW), weaning weight (WW), body weight at 6 months of age (W6) and yearling weight (YW). Records of 1,745 calves (1,513 from Hanwoo, and 232 from Korea Brindle calves) were collected from Livestock Research Institutes in Kangwon, Gyeongbuk and Chungbuk provinces. The least squares means (LSM) and their standard errors for BW, WW, W6 and YW were 25.4±0.1 kg, 81.0±1.8 kg, 146.1±3.7 kg and 291.5±2.4 kg, respectively in Hanwoo calves and 22.6±0.3 kg, 79.9±2.3 kg, 137.6±4.6 kg and 249.3±6.6 kg, respectively in Korea Brindle calves. Pedigree data showed that 14.8% (316 out of 2131) of Hanwoo was inbred and the average inbreeding coefficient was 0.0209 (2.09%). Inbreeding coefficients of ten calves out of 316 total inbred Hanwoo calves were 12.5% or higher, whereas those of the other 306 calves were less than 12.5%. In both breeds, calves were divided into three groups of inbreeding classes - highly inbred group(F≥ 0.125), lowly to medially inbred group(0<F<0.125) and no inbred group(F=0). In Korea Brindle calf populations, 12.2% of the calves observed (57 out of 467 calves) were inbred and the average inbreeding coefficient was 0.1367(13.67%). Forty four calves out of 57 inbred Korea Brindle calves had inbreeding coefficients of 12.5% or higher and the other 13 calves had less than 12.5% of inbreeding coefficients. Average inbreeding coefficient and the number of calves with greater than 12.5% inbreeding coefficient were higher in Korea Brindle calf groups than in Hanwoo calf groups. On the average, body weight growth of Korea Brindle calves was slower than that of Hanwoo calves. This would be due to very small breeding population structure of Korea Brindle cattle as compared to Hanwoo cattle, which would lead to rapid increase in inbreeding coefficients in the population. In conclusion, our study suggests that planned mating system is needed to control inbreeding in Korea Brindle population.
A total of 30 Korean native pigs (gilt 15, boar 15) were used to investigate the carcass characteristics, meat quality, amino acid, and fatty acid composition by gender. The carcass weight of boars were significantly higher than gilts, whereas the carcass yield of gilts had significantly higher than boars (p<0.01). Boars had significantly higher moisture contents in loin muscle than gilts, whereas the protein contents of loin muscle had significantly higher in gilts than boars (p<0.01). In the results of meat quality analysis, the cooking loss (p<0.01), shearing force (p<0.05), lightness (L) and yellowness (b) in meat color (p<0.05) were significantly higher, but the pH was significantly lower (p<0.01) in gilts compared with boars. Arginine (p<0.05), alanine, aspartic acid, histidine, leucine, lysine, phenylalanine, serine, threonine and tyrosin (p<0.01) for gilts were significantly higher than those for boars. The results of fatty acid composition showed that gilts had significantly higher contents of C16:1n7, C18:1n9, C20:1n9 (p<0.01) than boars in intramuscular fat, whereas boars had significantly higher contents of C18:2n6, C20:4n6 (p<0.01) and C18:3n3 (p<0.05) than gilts in intramuscular fat.
This study aimed to improve the reproductive efficiency of dairy herds by comparison and analyzing estrous appearance rate, conception and non-conception rate according to the stage of lactation using the lactation and reproductive records of average (less than 10,000 liters milk in 305 days) and high yielding (more than 10,000 liters milk in 305 days) Holstein cows (n=102). Milk production and reproduction data were collected between January 2010 and December 2012 from Holstein cows kept in the commercial dairy farms. Average (n=32) and high yielding (n=24) Holstein cows used to analyze the relationship between milk yield and reproductive performance. Our results showed that estrous appearance rate according to the stage of lactation was 25.0% (30∼59d), 40.6% (60∼ 89d), 25% (90∼110d) and 9.4% (>111d) in average yielding cows and 16.7% (30∼59d), 20.8% (60∼89d), 12.5% (90 ∼110d) and 50.0% (>111d) in high yielding cows, respectively. Conception rate according to the stage of lactation was 87.5% (30∼59d), 61.5% (60∼ 89d), 75.0% (90∼110d) and 66.7% (>111d) in average yielding cows and 25.0% (30∼59d), 0% (60∼89d), 33.3% (90∼ 110d) and 50.0% (>111d) in high yielding cows, respectively. Days between parturition and conception was 23.7% (<149d), 0% (150∼209d) and 0% (>210 d) in average yielding cows and 69.0% (<149 d), 77.8% (150∼209d) and 38.9% (>210d) in high yielding cows, respectively. Conception rate from 110 days postpartum in high yielding cows was 41.7% (110∼150d), 50.0% (151∼180d) and 50.0% (>181d). Body condition score (BCS) in 120 days postpartum was 2.64±0.1 in average yielding cows and 2.28±0.1 in high yielding cows, respectively.
Spermatogenesis is initiated from spermatogonial stem cells (SSCs) that has an ability of self-renewal and unipotency to generate differentiating germ cells. The objective of this study is to develop the simple method for derivation of SSCs using non-sorting of both spermatogonia and feeder cells. Simply uncapsulated mouse testes were treated with enzymes followed by surgical mincing, and single cells were cultured in stempro-34TM cell culture media at 37℃. After 5 days of culture, aciniform of SSC colony was observed, and showed a strong alkaline phosphatase activity. Molecular characterization of mouse SSCs showed that most of the mouse SSC markers such as integrin α6 and β1, CD9 and Stra8. In addition, pluripotency embryonic stem cell (ESC) marker Oct4 were expressed, however Sox2 expression was lowered. Interestingly, expression of SSC markers such as Vasa, Dazl and PLZF were stronger than mouse ESC (mESC). This data suggest that generated mouse SSCs (mSSCs) in this study has at least similar biomarkers expression to mESC and mSSCs derived from other study. Immunocytochemistry using whole mSSC colony also confirmed that mSSCs generated from this study expressed SSC specific biomarkers such as c-kit, Thy1, Vasa and Dazl. In conclusion, mSSCs from 5 days old mouse testes were successfully established without sorting of spermatogonia, and this cells expressed both mESC and SSC specific biomarkers. This simple derivation method for mSSCs may facilitate the study of spermatogenesis.
The aim of this study was to evaluate the changes of protein patterns in granulosa cells and corpus luteum in ovaries during the estrus cycle in cows. The estrus cycle was devided into five steps of follicular, ovulatory, early-luteal, mid-luteal and late-luteal phases. In results, 61 spots of total 85 spots were repeated on follicular phase and 51 spots of total 114 spots were repeated on ovulatory phase. The 40 spots of total 129 spots were repeated on early-luteal phase and 49 spots of total 104 spots were repeated on mid-luteal phase. Also 41 spots of total 60 spots were repeated on late-luteal phase. On the other hands, the 16 spots were indicated difference in follicular phase and ovulation phase had a difference 10 spots. It was showed difference No. 103 spot in ovulation phase, No. 135 spot in early-luteal phase and No. 175 and 176 spots in mid-luteal phase. Also, the 11 spots were expressed specifically in mid-luteal phase and No. 178 and 179 spots were difference of expression in late-luteal phase. We confirmed that there were 7 spots for ovulation, 4 spots for luteinization and 2 spots for luteolysis. Spot No. 89~93 in ovulation phase were transferrin, and spot No.94~98 were HSP60. Spot No. 103 was Dusty PK, spot No. 135 was OGDC- E2, and spot No. 175 and 176 were Rab GDI beta from luteinization. Spot No. 178 and 179 in luteolysis were vimentin. This results suggest that will be help to basic data about infertility.
This study was carried out to investigate synthetic extender for semen cryopreservation of Jeju Native Black Bull. The semen was collected using an artificial vagina and transported to the laboratory. The semen was diluted 1:1 by Tris-Egg yolk extender and contrifuged in 1,500 rpm for 15 minutes. The supernatant was removed. The pellect was diluted to final sperm concentration of 2×108/ml by doubling in every 30 minutes at 4℃ cold chamber. The semen was equilibrated for 4 hours at cold chamber and packed to 0.5 ml straw. The semen straws were located above 5 cm for 10 minutes. The height and duration affect the freezing speed by temperature. The frozen straw was plunged to LN2. The presented straws were examined the viability and motility after thawed at 37℃ water bath. Frozen-thawed sperm were evaluated sperm viability, membrane integrity and acrosome integrity. Post-thawed sperm viability has been significantly higher (p<0.05) in fresh sperm (93.27±1.62%) than frozen-thawed sperm (73.34±3.27%). However, there were no significant differences between fresh and frozen-thawed dead cell rate (7.35±2.63 vs, 13.71±2.85). In sperm motility, between Triladyl and AndroMed Extender, there was no significant different (72.86±2.83 vs, 81.47±2.48), similarly, the dead cell rates was similar (18.41±3.42% and 17.26±4.25). The results of our study suggest that AndroMed to the freezing extender showed more positive effect on the frozen-thawed spermatozoa in Jeju Native Black bull semen.
The karyotype of Korean short-hair cat was presented using the G-, C- and NOR- banding techniques. For chromosomes preparation, the fetus skin fibroblast cells were cultured and metaphases were obtained. In results, the Korean short-hair cat had 38 chromosomes with XX or XY, which consisted of 5 pairs of metacentric chromosomes(Group A and C), 3 pairs of submetacentric chromosomes (Group B), 6 pairs of medium metacentric chromosomes except for 1 pair of medium submetacentric D2 chromosomes (Group D, E), 2 pairs of acrocentric chromosomes(Group F) and metacentric X and Y sex chromosomes. In G-banding analysis, the Korean short-hair cat exhibited a typical and identical G-banding pattern in each homologous chromosome. Total number of bands and landmarks on the G-banded chromosomes of Korean short-hair cat well correspond to those of international standardization of karyotype of domestic cat. The heterochromatins of Korean short-hair cat chromosomes distributed at terminal and/or centromere regions on almost chromosomes by C-banding analysis. In addition, the C-banding pattern showed greatly heteromorphic in some chromosomes. Using the AgNOR-staining, we found the nucleolar organizer regions(NORs) of Korean short-hair cat located at chromosomes 1p12 site in E group. The quantity and number of NORs were constant among cells.