69 Cloned bovine embryonic development derived from interferon tau knockout cells

2019 ◽  
Vol 31 (1) ◽  
pp. 160 ◽  
Author(s):  
K.-M. Kim ◽  
S.-J. Lee ◽  
S.-Y. Yum ◽  
H.-S. Kim ◽  
H.-J. Kim ◽  
...  
Keyword(s):  
Embryonic Development ◽  
Formation Rate ◽  
Development Program ◽  
Donor Cell ◽  
Type I ◽  
Cleavage Rate ◽  
Interferon Tau ◽  
Signal Molecule ◽  
Significant Difference

Interferon tau (IFNT), a type I interferon, is known as a key signal molecule during pregnancy in ruminants because of the necessity in maternal recognition of pregnancy. It is produced in trophectoderm cells of the elongation bovine conceptus at Day 13-21, and peak production is at Day 15 to 17 of pregnancy. In addition, other studies show that it can affect embryonic development and quality. In this study, the effect of IFNT knockout in donor cells to bovine cloned embryonic development by somatic cell nuclear transfer (SCNT) was investigated. To proceed with this experiment, immature oocytes from ovaries at a local slaughterhouse were matured in vitro for 22h. To prepare the donor cell with IFNT knockout, somatic cells were transfected with Cas9 and single guide RNA targeting IFNT, and several single derived colonies with high proliferation were isolated for mutation assay. Finally, one colony that had mono-allelic mutation (4bp deletion) was selected and used as the donor cell for SCNT. A donor cell was injected into an enucleated oocyte. Reconstructed oocytes with the donor cell were fused by electrical shock, activated by chemical stimulation, and cultured for 7 days in chemically defined medium. For this study, control (n=94) and IFNT knockout groups (n=140) were compared with 4 replications. The results showed no significant difference between control and IFNT knockout groups not only in cleavage rate, but also in blastocyst formation rate (control: 15.7±8.3%, IFNT knockout group: 26.3±13.1%). In addition, the number of blastocyst cell was not different between control (88.2±27.0) and IFNT knockout group (88.0±21.1). Some IFNT mutated blastocysts from SCNT were randomly selected for confirmation of the deletion of IFNT, and all samples were positive for mutation. In conclusion, these data demonstrated that the disruption of IFNT did not affect embryonic development. In future study, we would transfer these embryos and check the effect of IFNT during pregnancy status. This work was supported by BK21 PLUS Program for Creative Veterinary Science, the National Research Foundation of Korea (2017R1A2B3004972), and the Technology Development Program (S2566872) by MSS.

154 THE EFFECT OF ZINC ON PORCINE IN VITRO MATURATION AND SUBSEQUENT EMBRYONIC DEVELOPMENT AFTER IN VITRO FERTILIZATION

2014 ◽  
Vol 26 (1) ◽  
pp. 191
Author(s):  
Y. Jeon ◽  
J. D. Yoon ◽  
L. Cai ◽  
S. U. Hwang ◽  
E. Kim ◽  
...  
Keyword(s):  
Embryonic Development ◽  
Inductively Coupled Plasma ◽  
In Vitro Maturation ◽  
Formation Rate ◽  
Developmental Competence ◽  
Developmental Potential ◽  
Vitro Fertilization ◽  
Nuclear Maturation ◽  
Significant Difference

Zinc (Zn) is one of the abundant transition metals in biology and is an essential component of most cells. However, there are few reports about the effect of Zn in porcine oocytes. The objective was to investigate the effects of supplementary Zn during in vitro maturation (IVM) of porcine oocytes. We investigated nuclear maturation, intracellular glutathione (GSH) levels, reactive oxygen species (ROS) levels, and subsequent embryonic development after IVF. Before the experiment, Zn concentrations in IVM medium and body fluids were measured using inductively coupled plasma spectrophotometer (sensitivity: 1 μM) and treatment concentrations were determined. Zinc concentration was 12.6 μM in porcine plasma and 12.9 μM in porcine follicular fluid. We confirmed that Zn was not detected in IVM medium. A total of 541 cumulus–oocyte complexes (COC) were used for the evaluation of nuclear maturation. The COC were matured in TCM-199 medium supplemented with various concentrations of Zn (0, 6, 12, 18, and 24 μM). After 44 h of IVM, no significant difference was observed in all groups (metaphase II rate: 85.7, 88.7, 90.4, 90.3, and 87.2%, respectively). A total of 100 matured oocytes were examined for the effects of different Zn concentrations (0, 6, 12, 18, and 24 μM) on porcine oocyte intracellular GSH and ROS levels, which were measured through fluorescent staining and image analysis program. The groups of 12, 18, and 24 μM showed a significant (P < 0.05) increase in intracellular GSH levels (1.45, 1.67, and 1.78, respectively) compared with the control and 6 μM group (1.00 and 1.08, respectively). The intracellular ROS level of oocytes matured with 12, 18, and 24 μM (0.82, 0.68, and 0.55) were significantly (P < 0.05) decreased compared with the control and 6 μM groups (1.00 and 1.03, respectively). Finally, the developmental competence of oocytes matured with different concentrations of Zn (0, 6, 12, 18, and 24 μM) was evaluated after IVF. There were no significantly different in cleavage rates. However, cleavage patterns and blastocyst (BL) formation were different. Fragmented embryo ratio of the 12 μM group (14.9%) was significantly lower than that of the other groups (control, 6, 18, and 24 μM: 26.4, 17.8, 18.4, and 18.0%, respectively). Oocytes treated with 12 μM Zn during IVM had a significantly higher BL formation rate (28.2%) after IVF compared with the control (19.8%). In conclusion, these results indicate that Zn treatment as body fluid concentration during IVM improved the developmental potential of IVF in porcine embryos by increasing the intracellular GSH concentration and decreasing the ROS level. This work was supported, in part, by a grant from the Next-Generation Bio Green 21 Program (No. PJ00956901), Rural Development Administration, and the National Research Foundation of Korea Grant funded by the Korean Government (NRF-2012R1A1A4A01004885, NRF-2013R1A2A2A04008751), Republic of Korea.

131 EFFECT OF BREED AND FROZEN - THAWED RAM SEMEN ON IN VITRO FERTILIZATION AND OVINE EMBRYONIC DEVELOPMENT

2011 ◽  
Vol 23 (1) ◽  
pp. 170 ◽  
Author(s):  
K. C. Lehloenya ◽  
N. Mahoete ◽  
J. P. C. Greyling ◽  
T. L. Nedambale
Keyword(s):  
In Vitro Fertilization ◽  
Embryonic Development ◽  
Developmental Stages ◽  
Germplasm Conservation ◽  
Egg Yolk ◽  
Blastocyst Formation ◽  
Cleavage Rate ◽  
Vitro Fertilization ◽  
Significant Difference

Ovine embryonic development was evaluated 8 days following in vitro fertilization, after using fresh or frozen–thawed Merino and indigenous (Pedi and Zulu) sheep semen. Semen used was collected twice weekly over a 3-month period with the aid of an electro-ejaculator. Following collection, semen samples were evaluated and semen with acceptable sperm motility and a percentage live sperm of 60% diluted with an egg yolk-based extender (Egg-Yolk Citrate). Semen samples were cryopreserved in straws with a programmable freezer to –130°C and then plunged into liquid nitrogen (–196°C) until used for IVF. Fresh and frozen–thawed semen was used to fertilize the matured oocytes in vitro. A total of 791 oocytes were fertilized using fresh semen and 802 oocytes fertilized using frozen–thawed semen. No significant differences were recorded between the fresh and frozen–thawed semen regarding the embryonic developmental stages. The performance of fresh and frozen–thawed semen followed the same trend, with the cleavage rate gradually declining with the progression in time and the embryonic developmental stage. The lowest developmental rate recorded was the occurrence of blastocyst formation, ranging between 0.4 ± 0.4% and 2.6 ± 1.0%. Regarding breed, no significant difference was observed from cleavage to the 2- to 4-cell stages. The use of fresh and frozen–thawed Zulu semen resulted in a significantly (P < 0.05) higher percentage of 8-cell development compared with the Pedi semen. However, the 8-cell embryonic stage recorded with the use of the Zulu ram semen (fresh and frozen–thawed), did not differ significantly from that of the Merino breed. No significant difference between the breeds regarding blastocyst formation was recorded. The overall cleavage rate, 2- to 4-cell, and blastocyst embryonic developmental stages following the use of fresh and frozen–thawed semen from the different rams were generally lower than those recorded by other researchers. The low blastocyst rates obtained warrant more research regarding the in vitro embryo production technique in order to improve the ovine blastocyst formation rate. The study was funded by the University of the Free State and conducted at the Germplasm Conservation and Reproduction Biotechnologies ARC.

189 THE EFFECTS OF RESVERATROL ON PORCINE OOCYTES IN VITRO MATURATION AND SUBSEQUENT DEVELOPMENT AFTER PARTHENOGENETIC ACTIVATION AND IN VITRO FERTILIZATION

2012 ◽  
Vol 24 (1) ◽  
pp. 207 ◽  
Author(s):  
S. S. Kwak ◽  
S. A. Jeong ◽  
Y. B. Jeon ◽  
S. H. Hyun
Keyword(s):  
Gene Expression ◽  
Embryonic Development ◽  
In Vitro Maturation ◽  
Formation Rate ◽  
Control Group ◽  
Developmental Potential ◽  
Parthenogenetic Activation ◽  
Nuclear Maturation ◽  
Significant Difference

The present study investigated the effects of resveratrol (a phytoalexin with various pharmacological activities) during in vitro maturation (IVM) of porcine oocytes on nuclear maturation, intracellular glutathione (GSH) and reactive oxygen species (ROS) levels, gene expression in matured oocytes and subsequent embryonic development after parthenogenetic activation (PA) and IVF. Data were analyzed with SPSS 17.0 using Duncan's multiple range test. In experiment 1, a total of 1146 cumulus–oocyte complexes (COC) were divided into 5 groups (0, 0.1, 0.5, 2.0 and 10.0 μM resveratrol). In the nuclear maturation after 44-h IVM, the groups of 0.1, 0.5 and 2.0 μM (83.0, 84.1 and 88.3%, respectively) had no significant difference compared to the control group (84.1%). The group of 10.0 μM decreased the nuclear maturation (75.0%) significantly (P < 0.05). In experiment 2, a total of 300 matured oocytes were examined for the effects of different resveratrol concentrations (0, 0.5, 2.0 and 10.0 μM) on porcine oocyte intracellular GSH and ROS levels. The groups of 0.5 and 2.0 μM showed a significant (P < 0.05) increase in intracellular GSH levels (1.3 and 1.3, respectively) compared with the control and 10.0 μM groups (1.0 and 1.0, respectively). The intracellular ROS level of oocytes matured with 2.0 μM resveratrol (0.4) was significantly (P < 0.05) decreased compared to other groups (control: 1.0; 0.5 μM: 0.6; and 10.0 μM: 0.7). In experiment 3, lower expression of apoptosis-related genes (Bax, Caspase-3 and Bak) was observed in matured oocytes treated with 2.0 μM resveratrol when compared with that of the control (P < 0.05). In experiment 4, a total of 728 oocytes were divided into 4 groups (control, 0.5, 2.0 and 10.0 μM) and examined subsequent to embryonic development after PA. Oocytes treated with 2.0 μM resveratrol during IVM had a significantly higher cleavage (CL) rate, blastocyst (BL) formation rate and total cell numbers (TCN) after PA compared with those of the control (2.0 μM: 96.6%, 62.1% and 49.1 vs control: 88.3%, 48.8% and 41.4, respectively) and the 10.0 μM groups (87.3%, 41.4% and 40.9, respectively). Oocytes treated with 0.5 μM resveratrol (87.2%, 50.5% and 48.6, respectively) during IVM had significantly higher TCN, but there were no differences in CL and BL formation rates. In experiment 5, a total of 935 oocytes in 3 groups (control, 2.0 and 10.0 μM resveratrol) were conducted in IVF. The BL formation rate and TCN were significantly higher in the group of 2.0 μM resveratrol (20.5% and 54.0, respectively) than the control (11.0% and 43.4, respectively) and 10.0 μM group (11.7% and 45.0, respectively), but there was no significant difference in CL rate. In conclusion, 2.0 μM resveratrol supplementation during IVM improved the developmental potential of PA and IVF in porcine embryos by increasing the intracellular GSH concentration, decreasing the ROS level and decreasing apoptosis-related gene expression during oocyte maturation. This work was supported by a grant from the Next-Generation BioGreen 21 Program (No. PJ008121), Rural Development Administration, Republic of Korea.

scholarly journals 299 IN VITRO DEVELOPMENT OF IMMATURE PORCINE OOCYTES FERTILIZED IN VITRO TO THE BLASTOCYST STAGE

2005 ◽  
Vol 17 (2) ◽  
pp. 300
Author(s):  
T. Somfai ◽  
K. Kikuchi ◽  
S.Y. Medvedev ◽  
A. Onishi ◽  
M. Iwamoto ◽  
...  
Keyword(s):  
Embryonic Development ◽  
Inner Cell Mass ◽  
Polar Body ◽  
Cell Mass ◽  
Cell Number ◽  
Blastocyst Stage ◽  
Oocyte Activation ◽  
Cleavage Rate ◽  
Significant Difference

In vitro fertilization (IVF) and embryonic development of mature and meiotically arrested porcine oocytes were compared in this study. After in vitro maturation (IVM) for 48 h of cumulus-oocyte complexes, 75.4% (n = 442) of them extruded a visible polar body (PB). Most oocytes with a polar body (PB+ group) were found to be at metaphase II (M-II) stage (91.4%). Most oocytes without a visible polar body (PB− group, n = 144) appeared to be arrested at the germinal vesicle (GV) (41.6%) and first meiotic metaphase (M-I) (34.0%) stages. After IVF of oocytes (the day of IVF = Day 0), there was no significant difference between PB+ and PB− groups in rates of sperm penetration, monospermy, and oocyte activation after the penetration. Embryonic development was assessed by staining with 1% orcein. On Day 2, although there was no difference between the embryo cleavage in PB+ (n = 447) and PB− (n = 217) groups (47.0% and 35.9%, respectively), PB+ embryos had more cells than the PB− embryos (3.37 and 2.81 cells, respectively) (P < 0.05; ANOVA). On Day 4, the cleavage rate of PB+ embryos was higher than that of PB− embryos (45.4% and 24.3%, respectively), and PB+ embryos had more cells than the PB− embryos (8.26 and 6.0 cells, respectively) (P < 0.05; ANOVA). On Day 6, a significantly higher number of PB+ embryos developed to the blastocyst stage than that of the PB− embryos (34.6% and 20.7%, respectively) (P < 0.05). However, by subtracting the GV oocytes from the PB− group, there was no difference in blastocyst rates between the M-I arrested and M-II oocytes (35.3% and 34.6%, respectively). The number of blastomer nuclei in embryos obtained from the PB+ group (52.0) was significantly higher than that of the PB− group (29.1); however, the proportion of inner cell mass and trophectoderm cells in PB+ and PB− blastocysts did not differ significantly (1:1.9 and 1:2.2, respectively) (P < 0.05). Chromosome analysis revealed that PB+ blastocysts had significantly more diploid blastomeres (69.7%) than PB− blastocysts (44.0%), whereas PB− blastocysts had significantly more triploid cells (34.0%) compared with PB+ oocytes (8.4%)(P < 0.05; χ2 test). These results indicate that porcine oocytes arrested at the M-I stage undergo cytoplasmic maturation during culture and have the same ability to develop to blastocysts after IVF as M-II oocytes but with a lower cell number; the latter might be caused by the slower embryonic development.

In vitro human masseter muscle hypersensitivity: a possible explanation for increase in masseter tone

1996 ◽  
Vol 80 (5) ◽  
pp. 1547-1553 ◽  
Author(s):  
P. J. Adnet ◽  
H. Reyford ◽  
B. M. Tavernier ◽  
T. Etchrivi ◽  
I. Krivosic ◽  
...  
Keyword(s):  
Masseter Muscle ◽  
Fiber Type ◽  
Vastus Lateralis ◽  
Threshold Concentration ◽  
Vastus Lateralis Muscle ◽  
Type I ◽  
Cross Sectional ◽  
Significant Difference ◽  
Maximal Tension

To determine whether a difference in fiber-type caffeine and Ca2+ sensitivities exists between human masseter and vastus lateralis skeletal muscle, we compared the fiber-type caffeine sensitivities in chemically skinned muscle fibers from 13 masseter and 18 vastus lateralis muscles. Caffeine sensitivity was defined as the threshold concentration inducing > 10% of the maximal tension obtained after the fiber was loaded with a 1.6 x 10(-2) mM Ca2+ solution for 30 s. Significant difference in the mean caffeine sensitivity was found between type I masseter fibers [2.57 +/- 1.32 (SD) mM] vs. type I (6.02 +/- 1.74 mM) and type II vastus lateralis fibers (11.25 +/- 3.13 mM). Maximal Ca(2+)-activated force per cross-sectional area was significantly different between masseter and vastus lateralis fibers. However, the Ca2+ concentration corresponding to half-maximal tension (pCa50) was not significantly different between type I masseter (pCa50 5.9 +/- 0.02) and type I vastus lateralis muscle (pCa50 6.01 +/- 0.08). These results suggest that the increase in caffeine sensitivity of masseter muscle reflects the presence of a low reactivity threshold of the sarcoplasmic reticulum.

scholarly journals 64COMPARISON OF DEVELOPMENTAL POTENTIAL OF IN VIVO AND IN VITRO RECIPIENT OOCYTES AFTER NUCLEAR TRANSFER IN GOAT

2004 ◽  
Vol 16 (2) ◽  
pp. 154
Author(s):  
H.S. Park ◽  
M.Y. Lee ◽  
S.P. Hong ◽  
J.I. Jin ◽  
J.K. Park ◽  
...  
Keyword(s):  
Nuclear Transfer ◽  
Electric Pulse ◽  
Serum Starvation ◽  
Cleavage Rate ◽  
Developmental Potential ◽  
In Vitro Development ◽  
Significant Difference ◽  
Vitro Development

Recent techniques in somatic cell nuclear transfer (SCNT) have been widely used for animal research. In addition, SCNT techniques may allow for the rescue of endangered species. Despite efforts for wildlife preservation, however, some threatened or endangered wild animal species will likely become extinct. As a preliminary experiment of a series in wildlife research, we tried to identify an improved method for the production of more transferable NT embryos in goats. Mature donor animals of Korean native goats (20–25kg) were synchronized with a CIDR (type G; InterAg, New Zealand) vaginal implant for 10 days followed by a total of 8 twice daily injections of 70mg of FSH (Folltropine, London, Ontario, Canada) and 400IU of hCG (Chorulon, Intervet, Moxmeer, The Netherlands). Oocytes were then collected surgically by retograde oviduct flush or direct aspiration from ovarian follicles in vivo at 29–34h after hCG. Oocytes collected from follicles were matured in TCM-199 containing 10% FBS and hormones. Prepared ear skin cells from the goat were cultured in TCM-199 containing 10% FBS at 39°C, 5% CO2 in air, and confluent monolayers were obtained. Oocytes were enucleated and donor cells from serum starvation (0.5%) culture were fused through a single electric pulse (DC 2.36kvcm−1, 17μs), and then activated by a single electric pulse (AC 5vmm−1, 5s+DC 1.56kvcm−1, 30μs) or chemical treatment (5μgmL−1 ionomycin 5min−1, 1.9mM 6-DMAP/4h). Reconstructed oocytes were cultured in M16 medium with 10% goat serum (GS) for 6–7 days. Data were analyzed by chi-square test. In in vitro development, significantly (P&lt;0.05) more oocytes were cleaved (24/30, 80.0%) and developed (7/24, 29.2%) to morula or blastocyst stage, respectively, in NT oocytes activated by Iono + DMAP compared to electric stimulated oocytes (2/21, 40.0%; 0/2, 0%). There was a significant difference in in vitro development of NT embryos by the method of oocyte collection. Cleavage rate was higher (P&lt;0.05) in NT embryos from in vivo oocytes (23/28, 82.1%) than in in vitro matured oocytes (19/35, 54.3%), and further development to morula or blastocyst was also significantly (P&lt;0.05%) higher in NT embryos from in vivo oocytes (7/23, 30.4%) than in NT embryos from in vitro matured oocytes (0/19, 0%). When we compared NT embryos to parthenotes, developmental rate was not significantly different between NT embryos and parthenotes. These results strongly suggest that the in vivo oocytes will have superior developmental potential to oocytes matured in vitro. Table 1 Effect of different oocyte source on in vitro development following caprine SCNT

277 FOLLICULAR GROWTH, SUBSEQUENT OVUM PICKUP, AND DOMINANT FOLLICLE REMOVAL IN COWS

2006 ◽  
Vol 18 (2) ◽  
pp. 246
Author(s):  
K. Imai ◽  
M. Tagawa ◽  
S. Matoba ◽  
M. Narita ◽  
K. Kanayama
Keyword(s):  
Formation Rate ◽  
Cumulus Cell ◽  
Oocyte Quality ◽  
Developmental Competence ◽  
Dominant Follicle ◽  
Cleavage Rate ◽  
Significant Difference ◽  
The Mean ◽  
Follicle Aspiration ◽  
Removal Group

The present study was designed to assess the recruitment of follicles after ovum pickup (OPU) and dominant follicle (DF) removal on the follicular wave after OPU in Holstein dry cows. Cows were reared under the same feeding and environmental conditions. In Experiment 1, follicle aspiration (>2 mm in diameter) by OPU using a 7.5-MHz linear transducer with needle (COVA needle; Misawa Medical, Tokyo, Japan) connected to an ultrasound scanner (SSD-1200; ALOKA, Tokyo, Japan) was performed in four cows. Then, ovaries were observed after OPU from Day 1 (Day 0 = the day of OPU) to Day 11 to assess the number of follicles developed. In Experiment 2, two sessions of OPU were performed with a 7 day interval between sessions, with or without dominant follicle removal, to assess the quality of developing follicles and oocytes. In the DF removal group, >8-mm follicles were aspirated at Day 5 after the first OPU session, and the same cows without DF removal were designated as a control (n = 4, crossover trial). Oocytes were evaluated by their cumulus cell morphology, cytoplasmic color, and density. To assess the developmental competence of oocytes, Grades 1 and 2 cumulus-oocyte complexes (COCs) were collected, matured, fertilized, and cultured as described by Imai et al. (2002 J. Vet. Med. Sci. 64(10), 887-891). Embryo development was assessed by the cleavage rate on Day 2 and the blastocyst formation rate on Days 7 to 9 (the day of insemination = Day 0). Data were analyzed by ANOVA or Student t-test. In Experiment 1, a dominant follicle (>8 mm in diameter) was developed during Days 3 to 5 after OPU in each donor. The mean number of developing follicles (>2 mm in diameter) were increased from Day 1 to Day 9 (Day 1: 7.5 � 2.1, Day 3: 19.0 � 1.2, Day 5: 23.3 � 9.0, Day 7: 30.3 � 11.0, Day 9: 42.0 � 15.8 and Day 11: 41.0 � 16.7 (mean � SD), P < 0.05). In Experiment 2, there was no difference in the mean number of developing follicles on the day of OPU and collected oocytes between DF removal and control groups (follicles: 47.8 � 23.0 and 39.3 � 6.2; oocytes: 27.0 � 11.6 and 26.5 � 5.4, respectively). The number of Grades 1 and 2 oocytes for the DF removal group was significantly higher (P < 0.05) than that for the control (83.6 � 1.5 and 63.2 � 14.2, respectively), and no significant difference was found within cleavage (60.0 � 37.2, 53.6 � 23.2) and blastocyst rates (34.1 � 33.9, 34.4 � 16.8). These results indicate that populations of follicles were increased till Day 9 after OPU, and the DF removal was effective at increasing oocyte quality in the developing follicles.

158 IMPACT OF MATURED CATTLE OOCYTES AT HIGHER INCUBATION TEMPERATURE ON IN VITRO EMBRYO PRODUCTION

2016 ◽  
Vol 28 (2) ◽  
pp. 209
Author(s):  
M. Nkadimeng ◽  
E. van Marle-Koster ◽  
K. P. M. Lekola ◽  
M. L. Mphaphathi ◽  
M. M. Seshoka ◽  
...  
Keyword(s):  
Incubation Temperature ◽  
Cell Types ◽  
Cell Number ◽  
Cleavage Rate ◽  
Embryo Production ◽  
Developmental Potential ◽  
Cell Numbers ◽  
Significant Difference ◽  
In Vitro Embryo Production

Heat stress during IVF is associated with reduced fertility in cattle oocytes. It may, however, enhance thermo-tolerance or cause detrimental effects on a variety of cell types or organisms, depending on the duration and intensity of the thermal challenge. The aim of this study was to evaluate the developmental potential of cumulus-oocyte complexes (COC) matured for 18 or 24 h and incubated at 39°C or 41°C. A total of 1000 immature oocytes were collected at slaughter from indigenous South African cow ovaries. The COC were randomly allocated (100/treatment) into 2 maturation times (18 or 24 h) and cultured in M199 + FSH-LH-estradiol medium under oil at 100% humidity and 5% CO2 at 39°C or 41°C. Post maturation, oocytes were subjected to normal subsequent embryo conditions. The Bracket and Oliphant medium was used for IVF. All matured oocytes were fertilised for 6 h with frozen-thawed Nguni bull semen at a concentration of 265 × 106. The presumptive zygotes from each treatment were cultured into SOF-BSA medium under oil and incubated at 39°C for assessment of cleavage rate 48 h post IVF. After Day 7 of culture, blastocyst were stained (Hoechst 33323) for nuclei cell count. Statistical analyses was performed using Genstat® software of SAS (SAS Institute, Cary, NC, USA; P < 0.05). Oocytes that were matured for 18 h in 41°C resulted in more 8-cell embryos (41%) compared with those incubated at 39°C (21.6%). However, no difference was observed for cleavage rate at both maturation times and incubation temperatures (41 or 39°C). There was more morula formation from oocytes matured for 18 h (19.6%) and 24 h (19.0%) at 41°C compared to 39°C (8.4%) group. The results further showed more blastocyst formation during 18 h at 41°C (15.2%) than at 39°C (7.4%) and during 24 h at 41°C (11.2%), 39°C (11.4%). However there was no difference in the nuclei cell number during 18 h at 41°C (45.2), 24 h (45.8), and 18 h at 39°C (43.4) of maturation. Thus, there was a significant difference in the nuclei cell numbers at 24 h on 39°C (n = 133.2) and 41°C (n = 45.8). In conclusion, oocytes that were matured for 18 and 24 h at 41°C or for 18 h at 39°C developed further to blastocyst stage on in vitro embryo production, however, with low nuclei cell numbers due to accelerated maturation temperature or shortened maturation period.

225 EFFECTS OF PEDIGREE, MEAT QUALITY, AND MEAT QUANTITY OF SLAUGHTERHOUSE DONOR COWS ON OOCYTE RECOVERY, EMBRYO DEVELOPMENT, AND PREGNANCY AFTER EMBRYO TRANSFER

2011 ◽  
Vol 23 (1) ◽  
pp. 211
Author(s):  
Y. S. Park ◽  
H. D. Park
Keyword(s):  
Pregnancy Rate ◽  
Meat Quality ◽  
Embryo Development ◽  
Development Program ◽  
Developmental Rate ◽  
Blastocyst Development ◽  
Oocyte Recovery ◽  
Significant Difference ◽  
Donor Cows

In conventional in vitro embryo production, oocytes are obtained from various ovaries, and donor characteristics, such as pedigree type, meat quality, or meat quantity, are not considered. The aim of this study was to examine the effects of pedigree type and meat quality and quantity of slaughterhouse donor cows on oocyte recovery, embryo development, and pregnancy rates. The ovaries of individual Korean native cows were obtained from a slaughterhouse. The ear tag numbers were obtained 24 h after slaughter, and the donor data were compared according to the pedigree type (no registration to third bloodline registration), meat quantity according to the percentage of carcass weight (more than 69%, 66 to 69%, and below 66%; Grades A, B, and C, respectively), and meat quality according to the marbling grade (marbling grade 8 to 9, 6 to 7, 4 to 5, 2 to 3, and 1; Grades 1++, 1+, 1, 2, and 3, respectively). There were 390 donors in total, and each group had 30 donors. Recipients (Holstein heifers; n = 222) were synchronized with a progesterone device, and a single blastocyst was transferred nonsurgically on Day 7 from the onset of oestrus. Data on embryo development were compared by Duncan’s multiple range tests, and pregnancy rate was analysed by chi-square test. Values of P < 0.05 were considered to indicate a significant difference. No significant difference was detected in the average number of oocytes recovered (17.5 ± 2.9 to 22.9 ± 1.2%) or blastocyst development (16.4 ± 2.6 to 19.5 ± 3.4%) from cows of different pedigrees. A higher number of oocytes were recovered from Grade A (22.9 ± 1.0) than from Grade B (19.8 ± 0.7; P < 0.05) cows. However, the meat quantity grade had no significant effect on blastocyst development (14.3 ± 1.2 to 15.0 ± 0.8%). A greater number of oocytes were recovered from the Grade 1++ (25.0 ± 1.6), 1+ (27.7 ± 2.4), and 2 (23.6 ± 1.5) groups than from the Grade 3 group (19.7 ± 1.0; P < 0.05). The developmental rate to the blastocyst stage was significantly higher in the Grades 2 and 3 groups (22.1 ± 2.2 and 19.4 ± 1.7) than in the Grade 1++, 1+, and 1 groups (9.0 ± 1.2 to 13.8 ± 2.0). Pregnancy rate was not affected by pedigree type, meat quantity, or meat quality (41.7 to 58.6%). We concluded that 1) pedigree type had no effect on oocyte recovery and embryo development; 2) meat quantity affected oocyte recovery but did not affect embryo development; 3) high meat quality (Grades 1++ and 1+) resulted in greater oocyte recovery but lower embryo development; and 4) pregnancy rate remained unaffected by pedigree type, meat quantity, and meat quality. This research was supported by the Technology Development Program for Agriculture and Forestry, Ministry for Food, Agriculture, Forestry and Fisheries, Republic of Korea.

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