scholarly journals Isolate® and Optiprep® minigradients as alternatives for sperm selection in bovine in vitro embryo production

2014 ◽  
Vol 94 (1) ◽  
pp. 35-42 ◽  
Author(s):  
L. L. Vianna ◽  
J. Pradieé ◽  
E. C. S. Santos ◽  
A. O. Gonçalves ◽  
L. F. M. Pfeifer ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Sperm Motility ◽  
Semen Quality ◽  
Membrane Integrity ◽  
Sperm Selection ◽  
Selection Methods ◽  
Embryo Production ◽  
Embryo Survival ◽  
In Vitro Embryo Production

Vianna, L. L., Pradieé, J., Santos, E. C. S., Gonçalves, A. O., Pfeifer, L. F. M., Rheingantz, M. G. T., Dode, M. A. N., Vieira, A. D., Lima, V. F. H., Correa, M. N. and Pegoraro, L. M. C. 2014. Isolate® and Optiprep® minigradients as alternatives for sperm selection in bovine in vitro embryo production. Can. J. Anim. Sci. 94: 35–42. The objective of this study was to evaluate alternatives in small volumes to conventional gradient of Percoll® on semen quality, in vitro embryo production, sex ratio and embryo survival after vitrification. Thawed semen was randomly allocated to one of four density gradient selection methods: (1) conventional Percoll® (P), (2) MiniPercoll (MP), (3) MiniIsolate (MI), and (4) MiniOptiprep (MO). Sperm kinetics and quality were evaluated. Use of P, MP and MI gradients did not affect sperm motility (P>0.05). However, there was a decrease in total and progressive sperm motility in MO (70.8 and 51.3% vs. 87.3 and 69.5% for P; 87.3 and 73% for MP; 92.3 and 78.8% for MI; P<0.05). The MO had lower membrane integrity compared with P, MP and MI (39.7 vs. 70.5, 72.3, 63.8%, respectively, P<0.05). The percentage of blastocysts produced was higher in MI than in MP and MO (21.1 vs. 16.1 and 16.9%, P<0.05) and similar to P (18.4%; P>0.05). Sex ratio and embryo survival after vitrification were similar among groups (P>0.05). Semen selected by Isolate and Optiprep gradient, at the concentrations and small volumes used, demonstrated similar characteristics and in vitro embryo production to conventional Percoll® gradient.

365 QUALITY ASSESSMENT OF BOVINE CRYOPRESERVED SPERM AFTER SEXING BY FLOW CYTOMETRY AND ITS USE FOR IN VITRO EMBRYO PRODUCTION

2010 ◽  
Vol 22 (1) ◽  
pp. 339
Author(s):  
J. O. Carvalho ◽  
R. Sartori ◽  
G. M. Machado ◽  
G. B. Mourão ◽  
M. A. N. Dode
Keyword(s):  
Flow Cytometry ◽  
Sperm Motility ◽  
Membrane Integrity ◽  
Fertilization Rate ◽  
Computer Assisted ◽  
Blastocyst Development ◽  
Embryo Production ◽  
Acridine Orange Staining ◽  
In Vitro Embryo Production

Several studies using sex-sorted sperm by flow cytometry have shown that its fertility is reduced. Therefore, this study evaluated structural and functional characteristics of sperm sexed by flow cytometry. In addition, in vitro embryo production (IVP) and development was assessed when frozen-thawed unsorted and sex-sorted sperm from 4 Nellore bulls. Each ejaculate was separated into three fractions: non-sexed (NS), sexed for X-sperm (SX), and sexed for Y-sperm (SY). After thawing, each sample was analyzed for sperm motility by computer-assisted semen analysis (CASA, Berkeley, CA), sperm head agglutination, sperm morphology, membrane integrity by propidium iodide (PI) and 6-carboxy-fluorescein diacetate (CFDA) staining, acrosome integrity by peanut agglutinin (PNA), capacitation by chlortetracycline (CTC), and chromatin integrity by acridine orange staining. Then, the samples were placed in 45 : 90% (NS90) or 45 : 60% (NS60, SX, and SY) Percoll™ gradients. After Percoll™ centrifugation, sperm pellets were analyzed or used for IVP. All analyses were replicated independently three times. For IVP, 2,271 in vitro matured oocytes were used. To assess fertilization rate, presumptive zygotes were fixed and stained with lacmoid at 18 h post-insemination (hpi). Cleavage was evaluated at Day 2 (48 hpi) and blastocyst development at Days 6, 7, 8, and 9 of culture. Data were analyzed using generalized linear models. No differences (P > 0.05) were observed between SX and SY groups for e sperm variables evaluated either before or after Percoll™. However, non-sexed sperm had higher sperm motility, greater percentage of sperm with intact membranes, and greater percentage of live sperm with intact acrosomes than sexed sperm (P < 0.05). An effect of Percoll™ was observed in the non-sexed samples, with those submitted to 45 : 90% gradient having higher motility, greater percentage of cells with intact membrane, and lower recovery rate than those submitted to a 45 : 60% gradient. No differences among groups were observed for fertilization rate, being 74.0 ± 5.7, 63.2 ± 5.1, 67.2 ± 5.7, and 55.4 ± 5.9% for NS90, NS60, SX, and SY, respectively. Group NS90 showed a greater cleavage rate than did the SY group, while groups NS60 and SX had similar rates to the others. Blastocyst development rates on Day 6 to Day 9 were greater for group NS90. For example, on Day 8 the blastocyst rate was 34.9 ± 3.6, 22.2 ± 3.2, 18.1 ± 3.3, and 14.8 ± 2.9% forNS90, NS60, SX, and SY groups, respectively. All groups showed similar embryonic developmental stages on Day 6 to Day 9. Although sex-sorting affected sperm characteristics, it did not cause a decrease with in vitro fertility. However, differences in blastocyst rates between groups NS60 and NS90 indicated that the sperm selection protocol affected embryo production. Financial support: Embrapa Genetic Resources and Biotechnology.

Relationship between the characteristics of frozen-thawed ram spermatozoa and in vitro embryo production

2001 ◽  
Vol 13 (3) ◽  
pp. 193 ◽  
Author(s):  
Lee H. A. Morris ◽  
W. H. Johnson ◽  
S. P. Leibo ◽  
B. C. Buckrell
Keyword(s):  
Logistic Regression ◽  
Sperm Motility ◽  
Membrane Integrity ◽  
Screening Tests ◽  
Breeding Programme ◽  
Embryo Production ◽  
Predictive Values ◽  
Commercial Breeding ◽  
In Vitro Embryo Production

To select rams suitable for ovine in vitro embryo production (IVP), the predictive values of the screening tests used to identify unsuitable rams need to be established. The present study examined some characteristics of frozen–thawed ram spermatozoa that might be evaluated routinely in a commercial breeding programme. These included sperm motility, plasma membrane integrity, morphology, and acrosome and capacitation status of the sperm population. Cryopreserved spermatozoa from four Dorset rams, which had previously satisfied the selection criteria for inclusion in a commercial breeding programme, were used for IVP. The overall contribution of the four rams and the ejaculates within each ram to the variability (R2) in the production of blastocysts was very small (2.1% and 2.5% respectively). The analysis of the sperm characteristics by logistic regression revealed a significant and positive association between total post-thaw sperm motility, viability and longevity with in vitro blastocyst production. However, there was no association between the other surface characteristics of the spermatozoa measured in this study with embryo production. Despite the absence of differences between the rams in the low incidence of polyspermic fertilization, the significant and detrimental effects of polyspermic fertilization on in vitro blastocyst production rates were quantified by logistic regression analysis. A large proportion of the variability within the IVP system was unaccounted for by the analysis of sperm and oocyte characteristics evaluated in this study. Thus, the identification of other factors contributing to the variability in the production of embryos in vitro warrants further investigation. No single sperm characteristic was sufficient to predict the ultimate outcome of blastocyst production. Rather, assessments of multiple characteristics within the IVP system are required to make accurate predictions.

scholarly journals Effect of different cryopreservation extenders added with antioxidants on semen quality and in vitro embryo production efficiency in cattle

2021 ◽  
Vol 93 (3) ◽  
Author(s):  
NATALIA C. SILVA ◽  
KAREN M. LEÃO ◽  
JOÃO T. PÁDUA ◽  
THAISA C. MARQUES ◽  
FRANCISCO R.A. NETO ◽  
...  
Keyword(s):  
Production Efficiency ◽  
Semen Quality ◽  
Embryo Production ◽  
In Vitro Embryo Production

180 ASSESSMENT OF BULL SEMEN QUALITY LOADED IN NEW SensiTemp STRAWS USING SEMEN AND IN VITRO PRODUCTION TECHNOLOGIES

2016 ◽  
Vol 28 (2) ◽  
pp. 221
Author(s):  
D. Le Bourhis ◽  
S. Camugli ◽  
P. Salvetti ◽  
L. Schibler ◽  
E. Schmitt
Keyword(s):  
Sperm Motility ◽  
Semen Quality ◽  
Sperm Quality ◽  
Semen Analysis ◽  
Membrane Integrity ◽  
Computer Assisted ◽  
Cleavage Rate ◽  
Fluid Medium ◽  
And Control

SensiTemp, a new in vitro maturation (IMV) bull straw concept, presents the advantage of colour changing while the straw is thawed. The colour of frozen straws is blue and straws start to become white when the temperature reaches 33°C, with a complete change of colour at 37°C. The objective of this study is to assess sperm quality after thawing of semen frozen in SensiTemp from 2 bulls, by analysing, in experiment 1, sperm motility and membrane integrity using computer-assisted semen analysis (CASA) and flow cytometry (FC), and, in experiment 2, the in vitro embryo production (IVP) using IVP technologies [IVM, IVF, and in vitro culture (IVC)]. The ejaculates of 2 bulls, selected during preliminary experiments on high in vitro fertility, were harvested at CIA L’Aigle, France, and split ejaculates were frozen in experimental (SensiTemp) and conventional (control) straws. In experiment 1 after thawing semen from the 2 types of straws (5 pooled straws each; 2 replicates), motility was assessed using the IVOS CASA system (Hamilton Thorne Inc., Beverly, MA, USA) and membrane integrity was evaluated through FC with Cytosoft software (Millipore-Guava Technologies Inc., Hayward, CA, USA). In experiment 2, IVF was used to evaluate the non-toxicity of SensiTemp and control straws. Cumulus-oocyte complexes (COC; n = 1178; 4 replicates) collected from slaughterhouse ovaries were matured in IVM medium (TCM-199 with bicarbonate, Sigma-Aldrich, Saint Quentin Fallavier, France; 10 µg mL–1 FSH-LH, Reprobiol, Liège, Belgium; and 10% FCS, Thermo Fisher, Illkirch, France) for 22 h. After fertilization, presumptive zygotes of each group (SensiTemp and control for each bull) were cultured in synthetic oviduct fluid medium (SOF, Minitube, Tiefenbach, Germany) with 1% estrous cow serum (ECS) and 0.6% BSA (Sigma-Aldrich, France) up to 8 days. All cultures were conducted at 38.5C in 5% CO2, and 5% O2. The cleavage and blastocysts rates were evaluated on Days 3 and 7, respectively, for each group. Embryo quality was recorded on Day 7 according to the IETS evaluation. Data from each bull were analysed separately using the chi-squared test (P < 0.05). In experiment 1, neither sperm motility from bull 1 (61.2 and 60.5%) and bull 2 (66.2 and 66.5%) nor membrane integrity from bull 1 (58.6 and 52.2%) and bull 2 (61.0 and 61.9%) were different between SensiTemp and control, respectively. Results from experiment 2 showed no difference (P > 0.05) in cleavage rate between SensiTemp and control for the 2 bulls: 92.1 and 91.7% for bull 1 and 94.2 and 94.6% for bull 2 respectively. The blastocysts rate on Day 7 did not differ (P > 0.05) among groups (47.5, 47.1 and 51.3, 50.4% for SensiTemp and control bull 1 and bull 2, respectively) nor the quality of embryos retrieved in the different groups: 25.4, 23.3, and 30.8, 29.6% in grade 1 embryo for SensiTemp and control bull 1 and bull 2, respectively. Those results demonstrate, in vitro, that the new SensiTemp straws were non-toxic and did not affect the semen quality after thawing nor did the SensiTemp straws affect the ability of sperm cells to fertilize oocytes and produce 8-day-old embryos.

18 EFFECTS OF SKIM MILK ON THE QUALITY AND FERTILITY OF BOAR SEMEN FOLLOWING LIQUID PRESERVATION AT 5°C AND 15°C

2011 ◽  
Vol 23 (1) ◽  
pp. 115 ◽  
Author(s):  
Z. Namula ◽  
R. Kodama ◽  
Y. Kaedei ◽  
F. Tanihara ◽  
V. L. Vien ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Storage Temperature ◽  
Semen Quality ◽  
Sperm Concentration ◽  
Membrane Integrity ◽  
Skim Milk ◽  
Control Group ◽  
Boar Semen ◽  
Boar Spermatozoa

Liquid preservation of semen can be an alternative to frozen–thawed semen for artificial insemination. The success of a selection of boar semen extenders has been studied over storage periods of 5 to 7 days. The objective of this study was to evaluate the effects of skim milk on the viability and in vitro fertility of boar spermatozoa preserved in Modena-based extenders at 5°C and 15°C for 2 weeks. A total of 7 ejaculates were collected from one boar. The sperm-rich fraction of each ejaculate was centrifuged and diluted in Modena extenders supplemented with 0 (control), 7.5, and 15 mg mL–1 of dry skim milk. The final sperm concentration was adjusted to 1 × 108 cells mL–1, and then the semen was stored at 5°C and 15°C for 2 weeks. In the first experiment, the motility, viability (live/dead fluorescence viability assay), plasma membrane integrity (hypoosmotic swelling test; HOST), and acrosome integrity (FITC-labelled peanut agglutinin staining) of semen stored for 2 weeks were assessed. In the second experiment, the fertilization of stored semen after 20 h of co-incubation with in vitro matured oocytes and their development were examined. Data were analysed using ANOVA. When the semen was stored at 5°C for 2 weeks, the mean total sperm motility of semen stored with 7.5 and 15 mg mL–1 of dry skim milk was significantly higher than that of semen in the control group (41.4% and 41.5% v. 17.4%; P < 0.05). However, the beneficial effects of skim milk on the sperm motility were not observed in the semen stored at 15°C. Moreover, there were no significant differences in the other parameters of semen quality among the groups in each storage temperature. Significantly higher penetration rates of semen stored with 7.5 and 15 mg mL–1 of dry skim milk were observed in the storage at 5°C (41.1% and 34.8% v. 19.8%; P < 0.05) but not at 15°C (38.9% and 26.0% v. 30.0%; P > 0.05) when compared with the control group. When the semen was stored at 5°C, the development rate to the blastocyst stage of oocytes fertilized with semen stored with 7.5 mg mL–1 of dry skim milk was significantly higher than that with control and 15 mg mL–1 of dry skim milk (15.4% v. 1.1% and 7.8%; P < 0.01). However, there were no significant differences in the development rates of oocytes fertilized with semen stored at 15°C among the groups (9.6–11.9%). In conclusion, our results indicate that the effect of skim milk on the viability and in vitro fertility of liquid-stored boar spermatozoa is dependent on the storage temperature. The addition of 7.5 mg mL–1 of dry skim milk may be effective for the improvement of viability and fertility of semen stored at 5°C but not at 15°C.

scholarly journals MicroRNA-210 Regulates Endoplasmic Reticulum Stress and Apoptosis in Porcine Embryos

Animals ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 221
Author(s):  
Muhammad Rosyid Ridlo ◽  
Eui Hyun Kim ◽  
Geon A. Kim
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Embryo Development ◽  
Caspase 3 ◽  
Formation Rate ◽  
Embryo Production ◽  
Embryo Survival ◽  
Expression Levels ◽  
In Vitro Embryo Production

Endoplasmic reticulum (ER) stress can be triggered during in vitro embryo production and is a major obstacle to embryo survival. MicroRNA (miR)-210 is associated with cellular adaptation to cellular stress and inflammation. An experiment was conducted to understand the effects of miR-210 on in vitro embryo development, ER stress, and apoptosis; to achieve this, miR-210 was microinjected into parthenogenetically activated embryos. Our results revealed that miR-210 inhibition significantly enhanced the cleavage rate, blastocyst formation rate, and total cell number (TCN) of blastocysts, and reduced expression levels of XBP1 (p < 0.05). miR-210 inhibition greatly reduced the expression of ER stress-related genes (uXBP1, sXBP1, ATF4, and PTPN1) and Caspase 3 and increased the levels of NANOG and SOX2 (p < 0.05). A miR-210-mimic significantly decreased the cleavage, blastocyst rate, TCN, and expression levels of XBP1 compared with other groups (p < 0.05). The miR-210-mimic impaired the expression levels of uXBP1, sXBP1, ATF4, PTPN1, and Caspase 3 and decreased the expression of NANOG and SOX2 (p < 0.05). In conclusion, miR-210 plays an essential role in porcine in vitro embryo development. Therefore, we suggest that miR-210 inhibition could alleviate ER stress and reduce apoptosis to support the enhancement of in vitro embryo production.

scholarly journals 240 GESTATION LENGTH AND BIRTH WEIGHT OF IN VITRO PRODUCED EMBRYOS FROM ZEBU DAIRY CATTLE

2005 ◽  
Vol 17 (2) ◽  
pp. 270 ◽  
Author(s):  
L.S.A. Camargo ◽  
J.H.M. Viana ◽  
A.A. Ramos ◽  
W.F. de Sa ◽  
A.M. Ferreira ◽  
...  
Keyword(s):  
Birth Weight ◽  
Sex Ratio ◽  
Sao Paulo ◽  
Control Group ◽  
Gestation Length ◽  
São Paulo ◽  
Embryo Production ◽  
In Vitro Embryo Production ◽  
And Control

Gir cattle are a popular zebu dairy breed in tropical and subtropical regions because of their tolerance of heat stress and resistance to tick-borne disease. The use of in vitro embryo production (IVP) technology may help accelerate genetic improvement of this breed. However, in general, IVP systems have been implicated in the production of large offspring and a greater proportion of male calves. Natural breeding results in newborn Gir calves weighing around 25 kg despite the fact that dams may weigh over 500 kg. It is unknown whether in vitro-produced Gir embryos also result in large offspring. The aim of this work was to evaluate the effect of in vitro embryo production on gestation length, birth weight, and sex ratio in Gir cattle. COCs were harvested by oocyte pickup from mature non-lactating Gir cows and in vitro-matured in TCM 199 medium (Gibco, Sao Paulo, Brazil) with 10% inactivated estrous cow serum for 24 h under 5% CO2 at 38.5°C in air. Gir spermatozoa were obtained through the swim-up method and co-incubated with oocytes in Fert-TALP media (Parrish JJ et al. 1988 Biol. Reprod. 38, 1171–1180) with 10 μg/mL heparin (Sigma, Sao Paulo, Brazil) and 6 mg/mL fatty acid-free bovine albumin (Sigma) for 18 h in 5% CO2 at 38.5°C in air. Presumptive zygotes were co-cultured with their own cumulus cells in CR2aa medium (Wilkinson RF et al. 1996 Theriogenology 45, 41–49) with 10% fetal calf serum in humid atmosphere of 5% CO2 at 38.5°C in air. Fresh Day 7 blastocysts were transferred to synchronized B. indicus × B. taurus crossbred recipients. Data of gestation length, birth weight, and gender ratio from 26 IVP calves (IVP group) were recorded and compared to data obtained from Gir calves produced by artificial insemination or natural mating (n = 24; control group) using ANOVA or chi-square analysis. There was no difference (P > 0.05) in gestation length between pregnancies of the IVP and control groups (mean ± SEM, 285.4 ± 1.5 vs. 284.4 ± 1.1 days, respectively). IVP and control calves showed similar (P > 0.05) weight at calving (29.6 ± 0.9 vs. 26.9 ± 1.2 kg for IVP and control male calves, and 27.0 ± 2.5 vs. 25.2 ± 0.5 kg for IVP and control female calves, respectively). The percentage of male calves was greater (P < 0.05) in the IVP group than in the control group (76.9% vs. 43.4%, respectively). IVP calves did not show abnormalities associated with Large Offspring Syndrome, such as breathing difficulty and perinatal death. These data suggest that in vitro production may affect the development of Gir embryos, biasing the sex ratio in a manner similar to previously reported for in vitro-produced embryos from Bos taurus breeds. This work was supported by FAPEMIG and CNPq.

scholarly journals In vitro embryo production in Nelore cows supplemented with calcium salts of long-chain fatty acid

2020 ◽  
Vol 41 (6) ◽  
pp. 2643-2654
Author(s):  
Moacir Rogério de Souza ◽  
◽  
Luciano Soares de Lima ◽  
Fabio Luiz Bim Cavalieri ◽  
Luiz Paulo Rigolon ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Soybean Oil ◽  
Pregnancy Rate ◽  
Fixed Time ◽  
Cleavage Rate ◽  
Embryo Production ◽  
Calcium Salts ◽  
In Vitro Embryo Production ◽  
Long Chain

This study was performed to evaluate the effects of supplementation of lactating Nelore cows with calcium salts of long-chain fatty acids (Ca-LCFA) of soybean oil on in vitro oocytes competency, cleavage rate, embryo production rate, pregnancy rate, and sex ratio of transferred embryos. Fifteen multiparous Nelore cows grazed on a Tifton 85 pasture were allotted to two treatments: (1) supplement without fat inclusion (CONT, n = 6) and (2) supplement containing 47.2 g Ca-LCFA /kg (to provide an intake of 0.2 kg Ca-LCFA/day) based on dry matter (n = 9). Animals were subjected to 2.53 ± 1.35 rounds of consecutive ovum pick up at intervals of 27.65 ± 21.72 days. Oocytes were morphologically evaluated, selected, maturated, fertilized, and cultivated for embryo production in vitro and then transferred into recipients who had previously undergone an embryo transfer protocol at a fixed time. The number of aspirated oocytes, the proportion of viable oocytes, cleavage rate, embryo percentage, pregnancy rate, and sex ratio were not affected when Nelore cows were supplemented with Ca-LCFA of soybean oil compared with non-supplemented Nelore cows. In conclusion, our results suggest that feeding a supplement containing 47.2 g Ca-LCFA/kg (to provide an intake of 0.2 kg Ca-LCFA/day) to Nelore cows does not affect in vitro embryo production in lactating Nelore cows. However, further studies using higher dietary doses of fat could be useful to determine the potential of fat supplementation to improve in vitro embryo production.

scholarly journals Laparoscopic Ovum Pick-Up Followed by In Vitro Embryo Production and Transfer in Assisted Breeding Programs for Ruminants

Animals ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 216
Author(s):  
Hernan Baldassarre
Keyword(s):  
Pregnancy Rate ◽  
Commercial Application ◽  
Embryo Production ◽  
Breeding Programs ◽  
Sheep And Goats ◽  
Embryo Recovery ◽  
Marker Selection ◽  
Holstein Calves ◽  
In Vitro Embryo Production

The potential of laparoscopic ovum pick-up (LOPU) followed by in vitro embryo production (IVEP) as a tool for accelerated genetic programs in ruminants is reviewed in this article. In sheep and goats, the LOPU-IVEP platform offers the possibility of producing more offspring from elite females, as the procedure is minimally invasive and can be repeated more times and more frequently in the same animals compared with conventional surgical embryo recovery. On average, ~10 and ~14 viable oocytes are recovered by LOPU from sheep and goats, respectively, which results in 3–5 transferable embryos and >50% pregnancy rate after transfer. LOPU-IVEP has also been applied to prepubertal ruminants of 2–6 months of age, including bovine and buffalo calves. In dairy cattle, the technology has gained momentum in the past few years stemming from the development of genetic marker selection that has allowed predicting the production phenotype of dairy females from shortly after birth. In Holstein calves, we obtained an average of ~22 viable oocytes and ~20% transferable blastocyst rate, followed by >50% pregnancy rate after transfer, declaring the platform ready for commercial application. The present and future of this technology are discussed with a focus on improvements and research needed.

Selection of stallions for in vitro embryo production by ICSI in a commercial program

2012 ◽  
Vol 32 (7) ◽  
pp. 409 ◽  
Author(s):  
C. Herrera ◽  
P. Dufourq ◽  
M. Freije ◽  
I. Morikawa ◽  
J.E. Centeno ◽  
...  
Keyword(s):  
Embryo Production ◽  
Commercial Program ◽  
In Vitro Embryo Production ◽  
Selection Of
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