232 ADDITION OF VERY LOW AMOUNTS OF SERUM (ESTRUS COW SERUM) IMPROVES IN VITRO EMBRYO PRODUCTION IN DAIRY CATTLE

2015 ◽  
Vol 27 (1) ◽  
pp. 205 ◽  
Author(s):  
E. Mullaart ◽  
F. Dotinga ◽  
C. Ponsart ◽  
H. Knijn ◽  
J. Schouten
Keyword(s):  
Culture Medium ◽  
Culture Media ◽  
Calf Serum ◽  
High Serum ◽  
In Vitro Production ◽  
Embryo Production ◽  
Chi Square ◽  
Embryo Yield ◽  
Vitro Production

Improving the efficiency of the in vitro production (IVP) process is very important because it results in more embryos to be used in breeding programs or as commercial service. At CRV, a culture medium consisting of SOF with amino acids and BSA is used. In the past, richer culture media were used with 10% fetal calf serum combined with BRL cell co-culture. Although the efficiency of the IVP process of these media was good, these rather high serum concentrations were quite often related to large offspring syndrome (LOS). The switch to a culture system without serum resulted in a significant reduction in LOS but also in a reduction of embryo yield. The aim of the present study was to investigate the effect of adding low amounts of serum to the culture medium on efficiency of embryo production. Immature cumulus-oocyte complexes (COC) were recovered from ovaries 6 to 8 h upon slaughter. The COC were matured in vitro in TCM199/FCS/LH/FSH supplemented with cysteamine (0.1 mM). Subsequently, matured oocytes were fertilised with frozen-thawed gradient-separated semen and further cultured for 7 days in SOFaaBSA. The SOF medium contained either 0 (control), 0.1, 0.5, or 1.0% oestrus cow serum (ECS). Embryos development was scored at Day 7. Three replicates were performed and results were analysed by chi-square analyses. The results clearly show that adding ECS significantly improved embryo production (Table 1). Interestingly, already very low amounts (0.1%) of serum gave a significant increase in embryo percentage. In conclusion, addition of very low amounts of ECS (0.1%) is beneficial for embryo production, resulting in significantly higher embryo production (from 19 to 27%). In a subsequent field trial with OPU-derived embryos, the effect of addition of 0.1% ECS on birth weight (LOS) of the calves has to be investigated. Table 1.Percentage of blastocysts at Day 7 after culture in SOF medium with different amounts of serum

Developments in in vitro technologies for swine embryo production

2004 ◽  
Vol 16 (2) ◽  
pp. 15 ◽  
Author(s):  
Matthew B. Wheeler ◽  
Sherrie G. Clark ◽  
David J. Beebe
Keyword(s):  
Culture Medium ◽  
Culture Media ◽  
Physical Culture ◽  
Efficient Production ◽  
Media Composition ◽  
In Vitro Production ◽  
Considerable Research ◽  
Blastocyst Rate ◽  
Vitro Production

Several modifications have been made to in vitro production (IVP) systems to allow more efficient production of viable porcine embryos. Although in vitro production of pig embryos has been studied for over 30 years, the overall blastocyst production rate remains low. The low blastocyst rate is due to several factors, including polyspermic oocyte penetration, low rate of male pronucleus formation and less than optimal in vitro culture systems. These conditions are all inherent problems in porcine IVP and many of the mechanisms involved remain unknown. Considerable research has examined culture medium and the techniques used during the various stages of in vitro production. However, changes to the physical culture system used during IVF have remained unchanged until recently. The present paper will summarise selected developments in fertilisation and embryo culture media composition and focus on the development of modified equipment to improve the conditions used during the IVP of porcine oocytes and embryos.

scholarly journals In vitro performance of Zebu (Bos indicus) and Taurus (Bos taurus) donor cow embryos

2020 ◽  
Vol 21 ◽  
Author(s):  
Anita Soares Barbosa GUIMARÃES ◽  
Laiara Fernandes ROCHA ◽  
Ronival Dias Lima de JESUS ◽  
Gisvani Lopes VASCONCELOS ◽  
Gabriela ANGHINONI ◽  
...  
Keyword(s):  
Culture Medium ◽  
Bos Taurus ◽  
Bos Indicus ◽  
In Vitro Production ◽  
Embryo Production ◽  
Maturation Medium ◽  
Significant Difference ◽  
Vitro Production ◽  
In Vitro Performance

ABSTRACT In this study, the in vitro production of bovine embryos from zebu and taurine donors was compared. Cumulus-oocyte complexes (COCs) were obtained from 167 Bos taurus and 161 Bos indicus donors by ovum pick-up. COCs were classified based on their morphological quality, matured in incubators for 22 to 24 h in maturation medium, and then fertilized for 18 to 22 h. The zygotes were transferred to the culture medium for seven days. The embryos were classified as morula (OM), initial blastocyst (BI), blastocyst (BL), and expanded blastocyst (BX), before being transferred to synchronized recipient cows. Pregnancy was diagnosed 30-45 days post-transfer. The Bos indicus donors had a higher oocyte yield (n = 2556) than Bos taurus donors (n = 1903) (P = 0.008). The COCs from zebu donors had a better morphological quality than those from taurine donors (n = 689 vs. 444 for grade 1 COC, P < 0.0001; n = 681 vs. 509 for grade 2 COC, P = 0.010, for zebu and taurine donors, respectively). There were differences in embryo production percentages obtained from OM (0.44% from zebu and 6.42% from taurine, P = 0.017), BL (14.18% from zebu and 3.74% from taurine, P < 0.0001), and BX (81.43% from zebu and 75.13% from taurine, P < 0.0001). No significant difference was observed for embryo production from BI and pregnancy rate (P > 0.05). The Bos indicus cows showed greater oocyte recovery, number of viable oocytes, and production of viable embryos than the Bos taurus cows.

scholarly journals 31 Effect of fetal calf serum on production and cryotolerance of in vitro bovine embryos from Ecuadorian Creole heifers

2019 ◽  
Vol 31 (1) ◽  
pp. 141
Author(s):  
M. S. Méndez ◽  
M. E. Soria ◽  
L. R. Galarza ◽  
F. P. Perea ◽  
D. E. Argudo
Keyword(s):  
Calf Serum ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Sodium Pyruvate ◽  
Embryo Production ◽  
Maturation Medium ◽  
Fetal Bovine ◽  
And Control ◽  
Vitro Production

In the Ecuadorian Andes there is a Creole bovine biotype whose population is disappearing. In vitro embryo production and cryopreservation is an important biotechnology that allows the conservation of animals threatened with extinction. The objective of this study was to determine the in vitro production and cryopreservation of embryos from creole heifers raised in the highlands of Ecuador. Immature cumulus-oocyte complexes were retrieved by ovum pickup from 10 Creole heifers (OPU) and from abattoir ovaries (control). The experiment was completed within 8 replicates. Cumulus-oocyte complexes were cultured in a maturation medium (TCM-199 supplemented with 10% fetal bovine serum, 100µg mL−1 of sodium pyruvate, 0.75mg mL−1 of l-glutamine, 4µg mL−1 of FSH-p, 100µM cysteamine, and 250µg mL−1 of gentamicin) following IVF (SOF medium supplemented with 10µg mL−1 heparin) and in vitro culture (citrate SOF medium). After denudation (Day 1 after IVF), presumptive embryos from each oocyte source (OPU and control) were split into 2 groups: with (FCS+) and without (FCS−) FCS (2.5%), which was added on Day 5 after IVF. On Day 7, embryos were evaluated, and those with quality 1 were vitrified. After warming, embryo re-expansion at 2h and embryo re-expansion and hatching at 24 and 48h were evaluated. Data were analysed by logistic regression in SAS software (SAS Institute Inc., Cary, NC, USA). Results of embryo rate at Day 7 and rates of vitrified, re-expanded, and hatched embryos are shown in Table 1. Regardless of the oocyte source, the addition of 2.5% FCS decreased embryo re-expansion at 2h and reduced embryo hatching at 48h in the OPU group. In conclusion, FCS did not improve embryo production and adversely affected the cryotolerance of embryos produced in vitro from Ecuadorian creole heifers. Table 1.Production and cryotolerance of in vitro bovine embryos

257 IN VITRO YAK EMBRYO PRODUCTION THROUGH CONVENTIONAL AND OVUM PICKUP METHODS

2015 ◽  
Vol 27 (1) ◽  
pp. 218
Author(s):  
P. Chakravarty ◽  
M. Hussain ◽  
M. S. Chauhan ◽  
R. S. Manik ◽  
D. Baishya ◽  
...  
Keyword(s):  
Conventional Method ◽  
Culture Medium ◽  
Slight Modification ◽  
In Vitro Production ◽  
Embryo Production ◽  
Maturation Medium ◽  
Vitro Production ◽  
Elite Germplasm ◽  
Successful Production

Yak is one of the most important economically useful animals for highlanders. The decline in the yak population demands effective measures for conservation and multiplication of elite germplasm. In vitro production of embryos and their cryopreservation and transfer to suitable recipients for production of elite calves may contribute to fulfill the objectives. The work was conducted at the National Research Center on Yak over a period of 3 years. The ovaries of slaughtered animals were used for collecting oocytes through aspiration of follicles followed by slicing of ovaries in the conventional method. Trials were conducted using 7 cyclic parous yaks for ultrasound-guided ovum pickup (OPU) at Nyukmadung farm (2700 m above mean sea level). The technique followed was similar to that in buffaloes with slight modification. Categories of oocytes classified A (2–3 layers of cumulus) and B (at least one layer of cumulus) obtained through the processes were subjected to in vitro maturation using standardized maturation medium (TCM-199 + 10% follicular fluid + sodium pyruvate + l-glutamine + 10% heat inactivated oestrus cow serum + pFSH + 17β oestradiol). The frozen-thawed yak sperm were capacitated using the swing-up method before their incubation with matured oocytes using BO medium. Oocytes matured for 24 h were washed 5 to 6 times with BO medium and then co-incubated with in vitro capacitated spermatozoa (0.1 to 0.25 million) for fertilization (8–10 oocytes per group) in 100-µL droplets of BO medium under mineral oil in 35-mm Petri dishes and placed in a CO2 incubator (5% CO2, 90% RH) at 38.5°C for 16 to 18 h. The presumed zygotes were washed several times in mCR2aa (modified Charles Rosenkrans) washing medium and then cultured in culture medium for 7 days on original beds of granulosa cells. The rates of maturation and fertilization of oocytes collected by conventional and OPU technique were comparable (Table 1). This may be attributed to greater numbers of good quality oocytes recovered in the conventional method. Embryos developed up to the stage of compact morula and blastocysts (24.66% through conventional and 22.73% through OPU) were cryopreserved using the vitrification method for further study. Thirteen embryos were transferred non-surgically to one each of 13 yak recipients; 5 became pregnant and only 1 recipient transferred with a cryopreserved-thawed embryo, developed through OPU, delivered one male calf, leading to the first successful production of an IVF yak calf in the world. The present findings are suggestive of using the OPU technique for in vitro embryo production, though resulting in lower numbers of transferable embryos (Table 1), because availability of ovaries for conventional IVF is a major constraint in yak. Table 1.Comparative in vitro yak embryo production rate with recovery of oocytes by conventional or ovum pickup (OPU) method

290 THE FACTORS ON RATES OF ABNORMALITY, DISEASE AND MORTALITY OF CALVES DERIVED FROM IN VITRO EMBRYOS OF KOREAN NATIVE CATTLE

2006 ◽  
Vol 18 (2) ◽  
pp. 252
Author(s):  
Y.-S. Park ◽  
S.-S. Kim ◽  
M.-C. Park ◽  
H.-D. Park
Keyword(s):  
Calf Serum ◽  
Treatment Technique ◽  
In Vitro Production ◽  
Chi Square ◽  
Offspring Number ◽  
Chi Square Test ◽  
Native Cattle ◽  
Vitro Production ◽  
Delivery Type

In Korea, in vitro production and transfer of bovine embryos has advanced remarkably and applied commercially. However, in vitro-produced embryos result in lower pregnancy and higher abortion rates and in some cases increased rates of abnormality and mortality in calves. The present study was conducted to investigate the effects of various factors such as recipient parity, delivery season, offspring number, pregnancy period, delivery type, midwifery type, dystocia and vaccination, on the viability of calves derived from embryos produced in vitro. Korean Native Cow ovaries were obtained from local slaughterhouse and cumulus-oocyte complexes (COCs) were aspirated from 2 to 8 mm follicles. Selected COCs were matured in TCM-199 supplemented with 10% fetal calf serum (FBS), 1 �ML FSH, 10 �ML LH and 1 �ML Estradiol-17� for 20-22 h. In vitro-matured oocytes were fertilized using frozen-thawed percoll separated semen (Day 0) in fer-TALP medium for 20 h. The presumptive zygotes were cultured in CR1aa medium supplemented with 0.3% BSA (before Day 3) or 10%FBS (After Day 3). All types of cultures were made in an incubator at 38.5�, 5% CO2 in air. Statistical analysis was performed using the Chi-square test. Two blastocysts were transferred to the Holstein recipients (n = 1888). The parturition was occurred in total 755 recipients. There was no difference in the abnormality of calves among treatments. The incidence of disease was significantly higher in single calf than twin calves (18.4 vs. 6.7%), in multiparous than nulliparous group (40.0 vs. 9.9%), in eutocia than dystocia group (20.0 vs. 4.8%), in spring and winter groups than summer and autumn groups (20.3, 22.7 vs. 4.3, 0.0%), and in non-vaccinated than vaccinated group (22.7 vs. 1.6%), respectively (p < 0.05). The rate of mortality was significantly higher when transferred into nulliparous than multiparous (22.3 vs. 0.0%), when were dystocia than eutocia group (71.4 vs. 14.1%), when were non-midwifery than midwifery (45.0 vs. 13.6), when delayed midwifery than earlier midwifery (31.6 vs. 11.5%), and when were non-vaccinated than vaccinated group (28.0 vs. 9.8%), respectively (P < 0.05). The present study suggested that the viability of bovine calves derived from in vitro was affected by the recipient parity, parturition treatment technique and vaccination. This study was supported by the BIO-GREEN 21 PROGRAM.

298 THE INFLUENCE OF DISTINCT CENTRAL BULL STATIONS ON THE IN VITRO EMBRYONIC DEVELOPMENT OF NELORE BULLS

2010 ◽  
Vol 22 (1) ◽  
pp. 305
Author(s):  
A. Renzi ◽  
F. P. Elias ◽  
R. A. Vila ◽  
R. B. Lôbo
Keyword(s):  
Calf Serum ◽  
Cumulus Cells ◽  
In Vitro Production ◽  
Freezing And Thawing ◽  
Chi Square ◽  
Frozen Semen ◽  
Chi Square Test ◽  
Vitro Production ◽  
In Vitro Embryonic Development

Reproductive biotechnology is growing worldwide as one of the most important tools of cattle breeding because it accelerates the process of genetic improvement. Most of the embryos produced are obtained using frozen semen from different AI centers. During freezing and thawing of semen, the sperm can be damaged by the rapid and dramatic changes in the physicochemical conditions that occur during cooling and ice formation. It has previously been described that bad management of frozen semen can result in reduced fertilization. This study investigated the influence of different central bull stations on the development of in vitro-produced bovine embryos. We compared semen of 154 Nelore bulls, used for IVF, from 8 different central bull stations (all of which used the same cryopreservation protocol) in the development of blastocysts. The in vitro production of embryos was performed as described: oocytes were collected from the slaughterhouse and matured in TCM-199 + 10% fetal calf serum (FCS) +0.5 μg mL-1 FSH + 50 μg mL-1 LH+ 1 μg mL-1 estradiol, for 24 hat 38.5°C in 5%CO2 in atmospheric air. Viable spermatozoids were obtained by centrifugation in Percoll gradient (45 and 90%), and used for IVF in a concentration of 2 million spermatozoa per mL in TALP + 10 μg mL-1 of heparin medium. After 12 h, the presumptive zygotes were transferred to a CR2+ 10% FCS medium and co-cultured with cumulus cells. After 168 h of IVF, we evaluated the number and stage of cleaved embryos produced with the semen of each bull. Statistical analyses were performed by using the chi-square test. Our results suggest that there are differences among distinct central bull stations in the proportion of embryos that developed into blastocysts and the different stages they hatched. FAPESP, CNPq, PROEX, FAEPA.

170 IN VITRO PRODUCTION OF BISON EMBRYOS

2012 ◽  
Vol 24 (1) ◽  
pp. 197 ◽  
Author(s):  
J. P. Barfield ◽  
G. E. Seidel
Keyword(s):  
Native American ◽  
Breeding Season ◽  
Culture Medium ◽  
Culture Media ◽  
Reproductive Technologies ◽  
In Vitro Production ◽  
Cell Stage ◽  
Maturation Medium ◽  
Vitro Production

The North American bison is a symbol of Native American heritage and the American West as well as being an increasingly important agricultural species. Reproductive technologies in bison lag behind those of cattle. Blastocyst production rates were low (<10%) in the only study of in vitro production of bison embryos (Thundathil et al. 2007 Theriogenology 68, 93–97). Our aims were to assess the application of our bovine in vitro embryo production system (De La Torre-Sanchez 2006 Reprod. Fertil. Dev. 18, 585–596) to bison gametes and evaluate the effect of adding FCS at different stages of the process. Initially, we performed homologous and heterologous inseminations with bison and cattle oocytes collected from abattoir ovaries and frozen-thawed epididymal bison sperm and ejaculated cattle sperm. Ovaries from both species were processed on the same day and a single straw of semen per species was used to fertilize all oocytes in each of 3 to 5 replicates. Culture media and conditions were identical for each treatment. Cleavage rates from the homologous IVF were 87% (86/99) for bison and 82% (164/199) for cattle. Bison oocytes with cattle sperm resulted in 88% (76/86) cleaved and cattle oocytes with bison sperm resulted in 69% cleaved (133/192; P < 0.01). Day 7 blastocyst rates per oocyte and per 8-cell stage, respectively, were 16 and 27% for cattle embryos, 6 and 13% for cattle oocytes with bison sperm, 10 and 16% for bison oocytes with cattle sperm and 7 and 9% for bison embryos. Experiments for bison embryos were then designed to evaluate the effect of adding 10% FCS to the maturation medium and 5, 2.5, or 0% to CDM2, the culture medium used after the 8-cell stage. Cleavage rates for oocytes matured in 10% FCS were 50% (202/405) and were 61% (206/337) in the absence of FCS. Day 7 blastocyst rates per oocyte and per 8-cell stage, respectively, of embryos matured in 10% FCS and then cultured in CDM2 + FCS were 12 and 26% in 5% FCS; 5 and 15% in 2.5% FCS; and 1 and 2.5% in 0% FCS. Similarly, Day 7 blastocyst rates of embryos matured without FCS but in CDM2 + FCS were 13 and 30% in 5% FCS, 1 and 4% in 2.5% FCS and 2 and 6% in 0% FCS. In a subsequent experiment, 5% FCS was added to CDM1 (culture medium for presumed zygotes through the 8-cell stage), CDM2, or both, but not to the maturation medium. Cleavage rates of bison embryos cultured with or without FCS in CDM1 were 63% (62/99) and 72% (145/202), respectively. Blastocyst rates of embryos cultured in CDM1 + FCS were 16% per oocyte and 36% per 8-cell stages in CDM2 + FCS and 0% in CDM2-FCS. Blastocyst rates of embryos cultured in CDM1 without FCS were 16% per oocyte and 25% per 8-cell stage in CDM2 + FCS and were 1% per oocyte and 2% per 8-cell stage in CDM2-FCS (P < 0.10). Adding 5% FCS to culture medium after embryos have reached the 8-cell stage greatly improved blastocyst rates of in vitro-produced bison embryos. Bison reproduction is highly seasonal and this work was conducted outside the bison breeding season. It is unknown if FCS would influence blastocyst rates when oocytes are collected from bison ovaries during the breeding season.

scholarly journals Cytokinin Mediated Increased In Vitro Production of Secondary Metabolites with Special Reference to Solasodine in Solanum erianthum

2021 ◽  
Vol 8 (02) ◽  
pp. e62-e68
Author(s):  
Jeeta Sarkar ◽  
Nirmalya Banerjee
Keyword(s):  
Secondary Metabolites ◽  
High Performance ◽  
Culture Medium ◽  
In Vitro Production ◽  
Leaf Extracts ◽  
Plant Parts ◽  
Regenerated Plants ◽  
Vitro Production ◽  
Regenerated Plantlets

AbstractSteroid alkaloid solasodine is a nitrogen analogue of diosgenin and has great importance in the production of steroidal medicines. Solanum erianthum D. Don (Solanaceae) is a good source of solasodine. The aim of this study was to evaluate the effect of different cytokinins on the production of secondary metabolites, especially solasodine in the in vitro culture of S. erianthum. For solasodine estimation, field-grown plant parts and in vitro tissues were extracted thrice and subjected to high-performance liquid Chromatography. Quantitative analysis of different secondary metabolites showed that the amount was higher in the in vitro regenerated plantlets compared to callus and field-grown plants. The present study critically evaluates the effect of the type of cytokinin used in the culture medium on solasodine accumulation in regenerated plants. The highest solasodine content (46.78±3.23 mg g-1) was recorded in leaf extracts of the in vitro grown plantlets in the presence of 6-γ,γ-dimethylallylamino purine in the culture medium and the content was 3.8-fold higher compared to the mother plant.

scholarly journals In Vitro Production of Erythropoietin by Kidneys Perfused With a Serum-free Solution

Blood ◽  
1974 ◽  
Vol 44 (1) ◽  
pp. 77-85 ◽  
Author(s):  
Allan J. Erslev
Keyword(s):  
Tissue Culture ◽  
Atmospheric Pressure ◽  
Culture Medium ◽  
Kidney Tissue ◽  
In Vitro Production ◽  
In Vitro Perfusion ◽  
Serum Free ◽  
Enzymatic Activation ◽  
Vitro Production

Abstract Normal rabbits exposed to 0.4 atmospheric pressure for 3 hr will generate about 40-60 U of erythropoietin during a subsequent 3-hr period. If the kidneys were removed from 3-hr hypoxic animals, washed carefully, and perfused for 3 hr by recirculation with a serum-tissue culture mixture, each kidney generated about 14 U of erythropoietin in vitro. Perfusion of normal kidneys did not result in the production of erythropoietin, and only small amounts were generated if the perfusate contained Puromycin. Three-hour hypoxic kidneys perfused for 3 hr with a serum-free tissue culture medium were found to generate about 8 U of erythropoietin per kidney and similar kidneys perfused with saline about 1 U. These results indicate that erythropoietin is synthesized by kidney tissue and not produced by enzymatic activation of a plasma substrate.

scholarly journals Fetal calf serum enhances in vitro production of Bos taurus indicus embryos

2011 ◽  
Vol 75 (3) ◽  
pp. 429-433 ◽  
Author(s):  
F.G. Leivas ◽  
D.S. Brum ◽  
S.S. Fialho ◽  
W.P. Saliba ◽  
M.T.T. Alvim ◽  
...  
Keyword(s):  
Fetal Calf Serum ◽  
Bos Taurus ◽  
Calf Serum ◽  
In Vitro Production ◽  
Bos Taurus Indicus ◽  
Vitro Production
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