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1.
Semina ciênc. agrar ; 42(1): 255-266, jan.-fev. 2021. tab
Artigo em Inglês | VETINDEX | ID: biblio-1501914

Resumo

The objective of this study was to evaluate the effect of and determine the optimum level of inclusion of docosahexaenoic acid (DHA) in the diluent for goat semen cryopreservation. Five Boer males underwent semen collection, totaling 10 viable collections per animal. After evaluation, the ejaculates were pooled and fractionated in Tris-yolk medium with the addition of 0; 30; 45; or 60ng mL-1 of DHA and 0.4 mmol of alpha-tocopherol (Vitamin E). The semen was cryopreserved in a freezing machine (TK 3000TM) and placed in a cryogenic cylinder for subsequent analysis. Data were evaluated by regression analysis at 5% significance. There were no differences (P > 0.05) in sperm kinetic parameters evaluated by computer assisted sperm analysis: total motility (79.17 ± 17.31%), progressive motility (14.04 ± 5.73%), curvilinear speed (58.82 ± 6.35µm/s), progressive linear speed (22.49 ± 3.63µm/s), mean path speed (35.17 ± 4.52µm/s), linearity (38.69 ± 5.79%), rectilinearity (63.99 ± 6.64%), and oscillation index (59.68 ± 2.99%). There were no differences (P > 0.05) found from the membrane functional integrity test for reactive spermatozoa (69.66 ± 9.76%), plasma and acrosomal membrane integrity of intact spermatozoa (29.86 ± 7.57%), mitochondrial potential of Class I cryopreserved goat semen (72.75 ± 9.81%), and chromatin compaction of intact chromatin (96.87 ± 4.37%).


O estudo teve como objetivo avaliar o efeito e determinar o melhor nível de inclusão de ácido docosahexaenoico (DHA) no diluidor para criopreservação de sêmen caprino. Cinco machos Boer foram submetidos a coletas de sêmen, totalizando 10 coletas viáveis por animal. Após avaliação, os ejaculados foram agrupados (pool) e fracionados em meio Tris gema, acrescido de 0; 30; 45 e 60ng mL-1 de DHA e 0,4mmol de alfa-tocoferol. O sêmen foi criopreservado em máquina de congelamento TK 3000® e acondicionado em botijão criogênico para posterior análise. Os dados foram avaliados por análise de regressão a 5% de significância. Não houve diferença (P > 0,05) nos parâmetros cinéticos espermáticos avaliados pela análise assistida por computador: motilidade total (79,17 ± 17,31%), motilidade progressiva (14,04 ± 5,73%), velocidade curvilínea (58,82 ± 6,35µm/s), velocidade linear progressiva (22,49 ± 3,63µm/s), velocidade média do caminho (35,17 ± 4,52µm/s), linearidade (38,69 ± 5,79%), retilinearidade (63,99 ± 6,64%) e índice de oscilação (59,68 ± 2,99%). Não foram encontradas diferenças (P> 0,05) no teste de integridade funcional da membrana para espermatozoides reativos (69,66 ± 9,76%), integridade plasmática e membrana acrossomal dos espermatozoides intactos (29,86 ± 7,57%), potencial mitocondrial do sêmen caprino de classe I (72,75 ± 9,81%) e compactação da cromatina intacta (96,87 ± 4,37%). A inclusão de até 60ng mL-1 de DHA não promoveu melhora nos parâmetros de qualidade seminal de caprinos pós-descongelamento.


Assuntos
Feminino , Animais , Análise do Sêmen/veterinária , Cabras/embriologia , Criopreservação/veterinária , Vitamina E/administração & dosagem , Ácidos Docosa-Hexaenoicos , /análise
2.
Semina Ci. agr. ; 42(1): 255-266, jan.-fev. 2021. tab
Artigo em Inglês | VETINDEX | ID: vti-31234

Resumo

The objective of this study was to evaluate the effect of and determine the optimum level of inclusion of docosahexaenoic acid (DHA) in the diluent for goat semen cryopreservation. Five Boer males underwent semen collection, totaling 10 viable collections per animal. After evaluation, the ejaculates were pooled and fractionated in Tris-yolk medium with the addition of 0; 30; 45; or 60ng mL-1 of DHA and 0.4 mmol of alpha-tocopherol (Vitamin E). The semen was cryopreserved in a freezing machine (TK 3000TM) and placed in a cryogenic cylinder for subsequent analysis. Data were evaluated by regression analysis at 5% significance. There were no differences (P > 0.05) in sperm kinetic parameters evaluated by computer assisted sperm analysis: total motility (79.17 ± 17.31%), progressive motility (14.04 ± 5.73%), curvilinear speed (58.82 ± 6.35µm/s), progressive linear speed (22.49 ± 3.63µm/s), mean path speed (35.17 ± 4.52µm/s), linearity (38.69 ± 5.79%), rectilinearity (63.99 ± 6.64%), and oscillation index (59.68 ± 2.99%). There were no differences (P > 0.05) found from the membrane functional integrity test for reactive spermatozoa (69.66 ± 9.76%), plasma and acrosomal membrane integrity of intact spermatozoa (29.86 ± 7.57%), mitochondrial potential of Class I cryopreserved goat semen (72.75 ± 9.81%), and chromatin compaction of intact chromatin (96.87 ± 4.37%).(AU)


O estudo teve como objetivo avaliar o efeito e determinar o melhor nível de inclusão de ácido docosahexaenoico (DHA) no diluidor para criopreservação de sêmen caprino. Cinco machos Boer foram submetidos a coletas de sêmen, totalizando 10 coletas viáveis por animal. Após avaliação, os ejaculados foram agrupados (pool) e fracionados em meio Tris gema, acrescido de 0; 30; 45 e 60ng mL-1 de DHA e 0,4mmol de alfa-tocoferol. O sêmen foi criopreservado em máquina de congelamento TK 3000® e acondicionado em botijão criogênico para posterior análise. Os dados foram avaliados por análise de regressão a 5% de significância. Não houve diferença (P > 0,05) nos parâmetros cinéticos espermáticos avaliados pela análise assistida por computador: motilidade total (79,17 ± 17,31%), motilidade progressiva (14,04 ± 5,73%), velocidade curvilínea (58,82 ± 6,35µm/s), velocidade linear progressiva (22,49 ± 3,63µm/s), velocidade média do caminho (35,17 ± 4,52µm/s), linearidade (38,69 ± 5,79%), retilinearidade (63,99 ± 6,64%) e índice de oscilação (59,68 ± 2,99%). Não foram encontradas diferenças (P> 0,05) no teste de integridade funcional da membrana para espermatozoides reativos (69,66 ± 9,76%), integridade plasmática e membrana acrossomal dos espermatozoides intactos (29,86 ± 7,57%), potencial mitocondrial do sêmen caprino de classe I (72,75 ± 9,81%) e compactação da cromatina intacta (96,87 ± 4,37%). A inclusão de até 60ng mL-1 de DHA não promoveu melhora nos parâmetros de qualidade seminal de caprinos pós-descongelamento.(AU)


Assuntos
Animais , Feminino , Cabras/embriologia , Criopreservação/veterinária , Análise do Sêmen/veterinária , Vitamina E/administração & dosagem , Ácidos Docosa-Hexaenoicos , Ácidos Graxos Ômega-3/análise
3.
Anim. Reprod. ; 18(4): e20200255, 2021. graf
Artigo em Inglês | VETINDEX | ID: vti-32761

Resumo

Here, we aimed to discriminate between the spectral profiles of spent culture media after oocyte in vitro maturation (IVM) and culture (IVC) from goats of different ages subjected to repeated hormonal treatments. The profiles were discriminated using near infrared (NIR) spectroscopy combined with multivariate methods. A total of 19 goats (young = 10; old = 9) were subjected to serial hormonal stimulation (HS) with gonadotropins. Cumulus oophorus complexes (COCs) were collected using laparoscopic ovum pick-up (LOPU) and subjected to IVM and parthenogenetic activation. The initial embryos were subjected to IVC. Spent culture media were collected after oocyte IVM and on day 2 of IVC and analyzed using NIR spectroscopy. NIR spectral data were interpreted through chemometric methods, such as principle component analysis (PCA) and partial least square discriminant analysis (PLS-DA). The results of PCA analysis clearly showed a separation in the spectral profiles between the experimental groups (HS sessions; young and old animals) both after IVM and IVC. Overall, the main absorption bands were attributed to the C-H group second overtone, first overtone of O-H and N-H, and C-H combinations and may serve as molecular markers. On the other hand, the spectral data obtained using PLS-DA models provided a better classification of the groups. The results showed the possibility of discriminating young and old groups as well as the three HS sessions with high specificity, sensitivity, and accuracy using NIR spectra. Thus, the culture medium analysis using NIR spectroscopy combined with multivariate methods indicated the dissimilarities between the groups and provided an insight into the in vitro development of goat oocytes. This technique serves as an efficient, objective, rapid, and non-invasive method to discriminate spectral profiles.(AU)


Assuntos
Animais , Feminino , Cabras/embriologia , Cabras/fisiologia , Análise Espectral , Técnicas de Maturação in Vitro de Oócitos
4.
R. bras. Reprod. Anim. ; 43(2): 235-241, abr.-jun. 2019.
Artigo em Português | VETINDEX | ID: vti-21842

Resumo

A redução da biodiversidade tem levado a uma preocupação emergencial frente a conservação do material biológico de animais geneticamente valiosos. Para tanto, a implantação de bancos de germoplasma, como embriões, gametas e tecido gonadal e somáticos, tem sido uma alternativa para auxiliar na manutenção da biodiversidade. No tocante ao material genético de fêmeas, a criopreservação de folículos pré-antrais isolados ou inclusos no tecido ovariano associada ao uso de tecnologias de reprodução assistida, como cultivo, fertilização in vitro e produção de embriões in vitro, poderá auxiliar na conservação da fertilidade de animais domésticos de interesse econômico com dificuldades reprodutivas ou animais ameaçados de extinção. Além disso, a criopreservação de tecido ovariano seguida de transplante pode ser uma alternativa promissora, uma vez que permite a restauração da função ovariana e reprodutiva. Estudos realizados em humanos relataram o nascimento de 130 indivíduos após o transplante de tecido ovariano previamente criopreservado. Apesar desse sucesso, a criopreservação de ovário ainda é considerada uma técnica experimental. Especialmente em caprinos e ovinos, a grande maioria dos estudos tem usado essas espécies como modelo experimental para a espécie humana. Nessas espécies, o sucesso da criopreservação de folículos ovarianos já foi evidenciado, o que nos dá esperança para o uso dessa tecnologia, vislumbrando, não somente a conservação do material genético, mas a obtenção de embriões e nascimento de crias mesmo após a morte de um animal geneticamente valioso. Assim, todos os esforços atuais estão voltados para a avaliação adequada da qualidade de protocolos de criopreservação que assegurem que esses recursos genéticos permaneçam como parte funcional dos sistemas de produção e conservação das espécies.(AU)


Reducing biodiversity has led to an emergency concern over the conservation of biological material from genetically valuable animals. Therefore, the establishment of gene banks, such as embryos, gametes and somatic and ovarian tissue has been an alternative to assist in maintaining biodiversity. Concerning the genetic material of females, the cryopreservation of preantral follicles isolated or included in the ovarian tissue associated with the use of assisted reproduction technologies, such as culture, in vitro fertilization and in vitro embryo production, may help to preserve the fertility of domestic animals of economic interest with reproductive difficulties or animals threatened with extinction. In addition, cryopreservation of ovarian tissue followed by transplantation may be a promising alternative, since it allows the restoration of ovarian and reproductive function. Studies in humans have reported the birth of 130 individuals after previously cryopreserved ovarian tissue transplantation. Despite this success, ovary cryopreservation is still considered an experimental technique. Especially in goats and sheep, the vast majority of studies have used these species as an experimental model for the human species. In these species, the success of cryopreservation of ovarian follicles has already been shown, which gives us hope for the use of this technology, noting not only the conservation of the genetic material, but the obtaining of embryos and the birth of youngsters even after the death of a genetically valuable animal. Thus, all current efforts are focused on the proper evaluation of the quality of cryopreservation protocols that ensure that these genetic resources remain as a functional part of the systems of production and conservation of the species.(AU)


Assuntos
Animais , Feminino , Criopreservação/veterinária , Fertilidade , Ovinos/embriologia , Ovinos/genética , Cabras/embriologia , Cabras/genética , Folículo Ovariano
5.
Anim. Reprod. ; 16(4): 819-828, 2019. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: vti-24100

Resumo

The objective of this study was to test the efficiency of powdered coconut water (ACP-406®) base-medium without or with the addition of supplements on in vitro culture of isolated goat secondary follicles. Follicles were cultured for 18 days in α-MEM or in ACP-406®, both without supplements (referred to as α-MEM and ACP, respectively), or both supplemented with BSA, insulin, transferrin, selenium, glutamine, hypoxanthine, and ascorbic acid (referred to as α-MEM+ and ACP+). Follicular morphology, antrum formation, follicular and oocyte diameter, levels of glutathione (GSH), and chromatin configuration after in vitro maturation were evaluated. At the end of culture, ACP-406® base-medium (without or with supplements) showed a higher (P < 0.05) percentage of normal follicles than α-MEM (without or with supplements). Antrum formation was similar among α-MEM+, ACP and ACP+, and significantly higher than α-MEM without supplements. The follicular diameter was greater in ACP+ than α-MEM, and similar to other treatments. Moreover, fully and daily grown rates were higher (P < 0.05) in ACP-406® base-medium (without or with supplements) than α-MEM (without or with supplements). Levels of GSH were similar between ACP+ and α-MEM+ treatments. Both ACP+ and α-MEM+ allowed meiotic resumption without a significant difference between the two groups. In conclusion, supplemented ACP-406® base-medium maintained follicular survival and promoted the development as well as meiotic resumption of isolated goat secondary follicles cultured in vitro for 18 days.(AU)


Assuntos
Animais , Feminino , Cabras/embriologia , Cabras/fisiologia , Folículo Ovariano/química , Folículo Ovariano/crescimento & desenvolvimento , Alimentos de Coco , Técnicas de Maturação in Vitro de Oócitos/veterinária
6.
Anim. Reprod. (Online) ; 16(4): 819-828, 2019. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461466

Resumo

The objective of this study was to test the efficiency of powdered coconut water (ACP-406®) base-medium without or with the addition of supplements on in vitro culture of isolated goat secondary follicles. Follicles were cultured for 18 days in α-MEM or in ACP-406®, both without supplements (referred to as α-MEM and ACP, respectively), or both supplemented with BSA, insulin, transferrin, selenium, glutamine, hypoxanthine, and ascorbic acid (referred to as α-MEM+ and ACP+). Follicular morphology, antrum formation, follicular and oocyte diameter, levels of glutathione (GSH), and chromatin configuration after in vitro maturation were evaluated. At the end of culture, ACP-406® base-medium (without or with supplements) showed a higher (P < 0.05) percentage of normal follicles than α-MEM (without or with supplements). Antrum formation was similar among α-MEM+, ACP and ACP+, and significantly higher than α-MEM without supplements. The follicular diameter was greater in ACP+ than α-MEM, and similar to other treatments. Moreover, fully and daily grown rates were higher (P < 0.05) in ACP-406® base-medium (without or with supplements) than α-MEM (without or with supplements). Levels of GSH were similar between ACP+ and α-MEM+ treatments. Both ACP+ and α-MEM+ allowed meiotic resumption without a significant difference between the two groups. In conclusion, supplemented ACP-406® base-medium maintained follicular survival and promoted the development as well as meiotic resumption of isolated goat secondary follicles cultured in vitro for 18 days.


Assuntos
Feminino , Animais , Alimentos de Coco , Cabras/embriologia , Cabras/fisiologia , Folículo Ovariano/crescimento & desenvolvimento , Folículo Ovariano/química , Técnicas de Maturação in Vitro de Oócitos/veterinária
7.
Rev. bras. reprod. anim ; 43(2): 235-241, abr.-jun. 2019.
Artigo em Português | VETINDEX | ID: biblio-1492574

Resumo

A redução da biodiversidade tem levado a uma preocupação emergencial frente a conservação do material biológico de animais geneticamente valiosos. Para tanto, a implantação de bancos de germoplasma, como embriões, gametas e tecido gonadal e somáticos, tem sido uma alternativa para auxiliar na manutenção da biodiversidade. No tocante ao material genético de fêmeas, a criopreservação de folículos pré-antrais isolados ou inclusos no tecido ovariano associada ao uso de tecnologias de reprodução assistida, como cultivo, fertilização in vitro e produção de embriões in vitro, poderá auxiliar na conservação da fertilidade de animais domésticos de interesse econômico com dificuldades reprodutivas ou animais ameaçados de extinção. Além disso, a criopreservação de tecido ovariano seguida de transplante pode ser uma alternativa promissora, uma vez que permite a restauração da função ovariana e reprodutiva. Estudos realizados em humanos relataram o nascimento de 130 indivíduos após o transplante de tecido ovariano previamente criopreservado. Apesar desse sucesso, a criopreservação de ovário ainda é considerada uma técnica experimental. Especialmente em caprinos e ovinos, a grande maioria dos estudos tem usado essas espécies como modelo experimental para a espécie humana. Nessas espécies, o sucesso da criopreservação de folículos ovarianos já foi evidenciado, o que nos dá esperança para o uso dessa tecnologia, vislumbrando, não somente a conservação do material genético, mas a obtenção de embriões e nascimento de crias mesmo após a morte de um animal geneticamente valioso. Assim, todos os esforços atuais estão voltados para a avaliação adequada da qualidade de protocolos de criopreservação que assegurem que esses recursos genéticos permaneçam como parte funcional dos sistemas de produção e conservação das espécies.


Reducing biodiversity has led to an emergency concern over the conservation of biological material from genetically valuable animals. Therefore, the establishment of gene banks, such as embryos, gametes and somatic and ovarian tissue has been an alternative to assist in maintaining biodiversity. Concerning the genetic material of females, the cryopreservation of preantral follicles isolated or included in the ovarian tissue associated with the use of assisted reproduction technologies, such as culture, in vitro fertilization and in vitro embryo production, may help to preserve the fertility of domestic animals of economic interest with reproductive difficulties or animals threatened with extinction. In addition, cryopreservation of ovarian tissue followed by transplantation may be a promising alternative, since it allows the restoration of ovarian and reproductive function. Studies in humans have reported the birth of 130 individuals after previously cryopreserved ovarian tissue transplantation. Despite this success, ovary cryopreservation is still considered an experimental technique. Especially in goats and sheep, the vast majority of studies have used these species as an experimental model for the human species. In these species, the success of cryopreservation of ovarian follicles has already been shown, which gives us hope for the use of this technology, noting not only the conservation of the genetic material, but the obtaining of embryos and the birth of youngsters even after the death of a genetically valuable animal. Thus, all current efforts are focused on the proper evaluation of the quality of cryopreservation protocols that ensure that these genetic resources remain as a functional part of the systems of production and conservation of the species.


Assuntos
Feminino , Animais , Cabras/embriologia , Cabras/genética , Criopreservação/veterinária , Fertilidade , Ovinos/embriologia , Ovinos/genética , Folículo Ovariano
8.
Semina Ci. agr. ; 40(4): 1501-1512, jul.-ago. 2019. tab
Artigo em Inglês | VETINDEX | ID: vti-21954

Resumo

The objective of this study was to evaluate the efficacy of delivering reduced doses of hormones via the Bai Hui acupoint in estrus synchronization in goats. A total of 40 goats received intravaginal sponges with medroxyprogesterone acetate for 7 days. The goats were then randomly distributed into 5 treatment: T1 - application of 132.5 μg of cloprostenol and 300 IU of equine chorionic gonadotropin (eCG), both by intramuscular injection (IM); T2 - application of 39.75 μg cloprostenol at the Bai Hui acupoint, and 300 IU of eCG by IM; T3 - application of 132.5 μg of cloprostenol by IM, and 90 IU of eCG at the Bai Hui acupoint; T4 - application of 39.75μg of cloprostenol and 90 UI of eCG, both in Bai Hui and T5 acupuncture: application of 39.75μg of cloprostenol and 90 UI of eCG, both applied in false acupoint. The goats were subjected to an estrus synchronization protocol and monitored for estrus detection, coverage and evaluation of reproductive parameters to detect entry into estrus. The data were subjected to normality tests, followed by appropriate statistical analyses of each variable. There was no significant difference (P > 0.05) in the percentage of animals in estrus (95.00 ± 11.18%), interval between sponge removal and beginning of estrus (49.72 ± 8.93 h), interval between sponge removal and end of estrus (76.84 ± 11.98 h), duration of estrus (27.08 ± 8.68 h), size of the largest follicle (6.82 ± 0.44 mm), interval between sponge removal and ovulation (78.28 ± 10.82 h), time from ovarian onset to estrus (28.52 ± 5.44 h), follicular growth rate (0.86 ± 0.29 mm/day), number of ovulations (1.32 ± 0.23), plasma progesterone concentration at 7 days after ovulation (10.28 ± 1.65 ng.mL-1), and gestation rate at 30 days after the beginning of estrus (75 ± 12.5%). However, the cost of the synchronization protocol per animal was 43.42% lower in treatments 4 and 5 (30% of the doses) than in treatment 1 (100% of the dose)...(AU)


O objetivo com o estudo foi avaliar a utilização de subdoses hormonais aplicadas no acuponto Bai Hui em protocolos de sincronização de estro em cabras. Foram utilizadas 40 cabras, as quais receberam esponjas intravaginais com acetato de medroxiprogesterona durante 7 dias, quando foram distribuídas aleatoriamente em 5 tratamentos: T1: aplicação de 132,5μg de cloprostenol e 300UI de eCG, ambos por via intramuscular (IM); T2: aplicação de 39,75μg de cloprostenol no acuponto Bai Hui e 300UI de eCG IM; T3: aplicação de 132,5μg de cloprostenol IM e 90UI de eCG no acuponto Bai Hui; T4: aplicação de 39,75μg de cloprostenol e 90UI de eCG, ambos no acuponto Bai Hui e T5: aplicação de 39,75μg de cloprostenol e 90UI de eCG, ambos aplicados em falso acuponto. As cabras foram submetidas à protocolo de sincronização de estro e monitoradas para detecção de estro, realização das coberturas e avaliação dos parâmetros reprodutivos. Os dados foram submetidos à análise de normalidade, seguido dos testes estatísticos adequados para cada variável. Não obteve-se diferença (p > 0,05) para porcentagem de animais em estro (95,00 ± 11,18%), intervalo da retirada da esponja ao início do estro (49,72 ± 8,93h), intervalo da retirada da esponja ao final do estro (76,84 ± 11,98h), duração do estro (27,08 ± 8,68h), tamanho do maior folículo (6,82 ± 0,44mm), intervalo da retirada da esponja à ovulação (78,28 ± 10,82h), intervalo do início do estro à ovulação (28,52 ± 5,44h), taxa de crescimento folicular (0,86 ± 0,29mm/dia), número de ovulações (1,32 ± 0,23), concentração de progesterona plasmática aos 7 dias após ovulação (10,28 ± 1,65ng.mL-1) e para taxa de gestação aos 30 dias (75 ± 12,5%). Porém, o custo do protocolo de sincronização por animal foi 43,42% menor para os tratamentos T4 e T5 (30% da dose) comparado com o T1 (100% da dose)...(AU)


Assuntos
Animais , Feminino , Cabras/embriologia , Terapia por Acupuntura/veterinária , Terapia por Acupuntura/métodos , Técnicas Reprodutivas/veterinária , Cruzamento/métodos , Sincronização do Estro/métodos
9.
Anim. Reprod. ; 16(4): 838-845, 2019. tab, graf
Artigo em Inglês | VETINDEX | ID: vti-24136

Resumo

This study evaluated a powdered coconut water solution (ACP 406®) as a base culture medium on the in vitro survival and development of in situ goat preantral follicles. The ovarian fragments were either immediately fixed in Carnoy solution (non-cultured control) or individually cultured for 2 or 6 days. The following culture media (all containing 100 μg/mL penicillin and 100 μg/mL streptomycin) were evaluated: α-MEM (α-MEM alone, without additional supplementation); α-MEM+ (supplemented α-MEM); ACP (ACP®406 alone); or ACP+ (supplemented ACP®406). Additional supplementation includes: 1.25 mg/mL bovine serum albumin, 10 μg/mL insulin, 5.5 μg/mL transferrin, 5 ng/mL selenium, 2 mM glutamine, and 2 mM hypoxanthine. The endpoints (i) follicular morphology; (ii) development; (iii) estradiol production; and (iv) reactive oxygen species (ROS) were recorded. Data were analyzed using chi-square, Turkey, t-test or One-Way ANOVA. Differences were considered significant when P < 0.05. At day 2 of culture, a greater (P < 0.05) percentage of morphologically normal follicles was observed between ACP+ and ACP treatments. Moreover, at day 2 of culture, no hormonal difference (P < 0.05) was observed between ACP+ and both α-MEM treatments. At day 6 of culture when ACP and α-MEM treatments were compared the percentage of healthy follicles were similar (P > 0.05) among treatments. Overall, all treatments had lower primordial follicles (P < 0.05) accompany by greater developing follicles (P < 0.05) percentages than non-cultured control treatment, indicating primordial follicle activation. However, at day 6 of culture, the percentage of primordial follicle development were similar (P > 0.05) among the treatments. Likewise, no differences (P > 0.05) were observed for ROS production and follicular and oocyte diameters among treatments. Therefore, ACP+ has the equivalent efficiency to MEM+ in maintaining the survival and development of goat preantral follicles, representing an alternative plant-based low-cost culture medium for in vitro culture.(AU)


Assuntos
Animais , Feminino , Cabras/embriologia , Alimentos de Coco , Fertilização in vitro/instrumentação , Fertilização in vitro/veterinária , Folículo Ovariano/crescimento & desenvolvimento
10.
Semina ciênc. agrar ; 40(4): 1501-1512, jul.-ago. 2019. tab
Artigo em Inglês | VETINDEX | ID: biblio-1501440

Resumo

The objective of this study was to evaluate the efficacy of delivering reduced doses of hormones via the Bai Hui acupoint in estrus synchronization in goats. A total of 40 goats received intravaginal sponges with medroxyprogesterone acetate for 7 days. The goats were then randomly distributed into 5 treatment: T1 - application of 132.5 μg of cloprostenol and 300 IU of equine chorionic gonadotropin (eCG), both by intramuscular injection (IM); T2 - application of 39.75 μg cloprostenol at the Bai Hui acupoint, and 300 IU of eCG by IM; T3 - application of 132.5 μg of cloprostenol by IM, and 90 IU of eCG at the Bai Hui acupoint; T4 - application of 39.75μg of cloprostenol and 90 UI of eCG, both in Bai Hui and T5 acupuncture: application of 39.75μg of cloprostenol and 90 UI of eCG, both applied in false acupoint. The goats were subjected to an estrus synchronization protocol and monitored for estrus detection, coverage and evaluation of reproductive parameters to detect entry into estrus. The data were subjected to normality tests, followed by appropriate statistical analyses of each variable. There was no significant difference (P > 0.05) in the percentage of animals in estrus (95.00 ± 11.18%), interval between sponge removal and beginning of estrus (49.72 ± 8.93 h), interval between sponge removal and end of estrus (76.84 ± 11.98 h), duration of estrus (27.08 ± 8.68 h), size of the largest follicle (6.82 ± 0.44 mm), interval between sponge removal and ovulation (78.28 ± 10.82 h), time from ovarian onset to estrus (28.52 ± 5.44 h), follicular growth rate (0.86 ± 0.29 mm/day), number of ovulations (1.32 ± 0.23), plasma progesterone concentration at 7 days after ovulation (10.28 ± 1.65 ng.mL-1), and gestation rate at 30 days after the beginning of estrus (75 ± 12.5%). However, the cost of the synchronization protocol per animal was 43.42% lower in treatments 4 and 5 (30% of the doses) than in treatment 1 (100% of the dose)...


O objetivo com o estudo foi avaliar a utilização de subdoses hormonais aplicadas no acuponto Bai Hui em protocolos de sincronização de estro em cabras. Foram utilizadas 40 cabras, as quais receberam esponjas intravaginais com acetato de medroxiprogesterona durante 7 dias, quando foram distribuídas aleatoriamente em 5 tratamentos: T1: aplicação de 132,5μg de cloprostenol e 300UI de eCG, ambos por via intramuscular (IM); T2: aplicação de 39,75μg de cloprostenol no acuponto Bai Hui e 300UI de eCG IM; T3: aplicação de 132,5μg de cloprostenol IM e 90UI de eCG no acuponto Bai Hui; T4: aplicação de 39,75μg de cloprostenol e 90UI de eCG, ambos no acuponto Bai Hui e T5: aplicação de 39,75μg de cloprostenol e 90UI de eCG, ambos aplicados em falso acuponto. As cabras foram submetidas à protocolo de sincronização de estro e monitoradas para detecção de estro, realização das coberturas e avaliação dos parâmetros reprodutivos. Os dados foram submetidos à análise de normalidade, seguido dos testes estatísticos adequados para cada variável. Não obteve-se diferença (p > 0,05) para porcentagem de animais em estro (95,00 ± 11,18%), intervalo da retirada da esponja ao início do estro (49,72 ± 8,93h), intervalo da retirada da esponja ao final do estro (76,84 ± 11,98h), duração do estro (27,08 ± 8,68h), tamanho do maior folículo (6,82 ± 0,44mm), intervalo da retirada da esponja à ovulação (78,28 ± 10,82h), intervalo do início do estro à ovulação (28,52 ± 5,44h), taxa de crescimento folicular (0,86 ± 0,29mm/dia), número de ovulações (1,32 ± 0,23), concentração de progesterona plasmática aos 7 dias após ovulação (10,28 ± 1,65ng.mL-1) e para taxa de gestação aos 30 dias (75 ± 12,5%). Porém, o custo do protocolo de sincronização por animal foi 43,42% menor para os tratamentos T4 e T5 (30% da dose) comparado com o T1 (100% da dose)...


Assuntos
Feminino , Animais , Cabras/embriologia , Cruzamento/métodos , Sincronização do Estro/métodos , Terapia por Acupuntura/métodos , Terapia por Acupuntura/veterinária , Técnicas Reprodutivas/veterinária
11.
Anim. Reprod. (Online) ; 16(4): 838-845, 2019. tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461468

Resumo

This study evaluated a powdered coconut water solution (ACP 406®) as a base culture medium on the in vitro survival and development of in situ goat preantral follicles. The ovarian fragments were either immediately fixed in Carnoy solution (non-cultured control) or individually cultured for 2 or 6 days. The following culture media (all containing 100 μg/mL penicillin and 100 μg/mL streptomycin) were evaluated: α-MEM (α-MEM alone, without additional supplementation); α-MEM+ (supplemented α-MEM); ACP (ACP®406 alone); or ACP+ (supplemented ACP®406). Additional supplementation includes: 1.25 mg/mL bovine serum albumin, 10 μg/mL insulin, 5.5 μg/mL transferrin, 5 ng/mL selenium, 2 mM glutamine, and 2 mM hypoxanthine. The endpoints (i) follicular morphology; (ii) development; (iii) estradiol production; and (iv) reactive oxygen species (ROS) were recorded. Data were analyzed using chi-square, Turkey, t-test or One-Way ANOVA. Differences were considered significant when P 0.05) among treatments. Overall, all treatments had lower primordial follicles (P 0.05) among the treatments. Likewise, no differences (P > 0.05) were observed for ROS production and follicular and oocyte diameters among treatments. Therefore, ACP+ has the equivalent efficiency to MEM+ in maintaining the survival and development of goat preantral follicles, representing an alternative plant-based low-cost culture medium for in vitro culture.


Assuntos
Feminino , Animais , Alimentos de Coco , Cabras/embriologia , Fertilização in vitro/instrumentação , Fertilização in vitro/veterinária , Folículo Ovariano/crescimento & desenvolvimento
12.
R. bras. Ci. Vet. ; 26(1): 21-25, jan.-mar. 2019. ilus, tab
Artigo em Inglês | VETINDEX | ID: vti-23090

Resumo

Objetivou-se descrever os distúrbios reprodutivos associados à infecção experimental por Toxoplasma gondii através da inseminação artificial com sêmen contaminado em quatro cabras no estágio crônico da infecção. As características do trato reprodutor foram avaliadas através de ultrassonografia transretal, visando o diagnóstico gestacional ou de desordens reprodutivas, após a infecção experimental. Ao final do experimento, os animais foram necropsiados e avaliações histopatológicas e PCR foram realizados. Dentre os animais infectados que exibiram mortalidade embrionária, duas apresentaram anestro e duas apresentaram repetição de estro, sendo que destas uma apresentou intervalos entre estros reduzido (sete dias) e outra em intervalo regular (21 dias). Todavia, ambas foram submetidas a monta natural durante os estros naturais subsequentes e não foi confirmada gestação até o final do experimento (90 dias). Duas cabras exibiram alterações nos exames de ultrassonografia, sendo identificadas um cisto ovariano, e uma hidrossalpinge, ambas confirmadas no exame post-mortem. As principais lesões microscópicas nesse grupo foram infiltração neutrofílica dos pulmões, glomerulonefrite intersticial e infiltração neutrofílica do fígado. O DNA de T. gondii foi encontrado nos órgãos (coração e cérebro) de três cabras. Em conclusão, cabras infectadas com sêmen contendo T. gondii no momento da inseminação artificial apresentam distúrbios reprodutivos na fase crônica da infecção que podem estar associados à toxoplasmose.(AU)


The aim of this study was to describe the reproductive disorders related to experimental infection by artificial insemination with semen contaminated with Toxoplasma gondii of four goats in the chronic phase of the infection. In the end of the study, the does were submitted to necropsy, and PCR and histopathological evaluations were performed. Among infected does that exhibited embryonic loss, two were in anestrus and two exhibited repeated estrus. One of the latter animals exhibited clinical signs of estrus at seven-day intervals, whereas the other had a 21-day estrous cycle. However, both does were naturally mated on subsequent natural estrous and were not able to get pregnant until the end of the experiment (90 d). Two of the goats exhibited abnormalities in the ultrasound examinations, one of which was an ovarian cyst, while the other was a hydrosalpinx, both of which were confirmed in the post-mortem examination. The main microscopic injuries in this group were neutrophilic infiltration of the lungs, interstitial glomerulonephritis and neutrophilic infiltration of the liver. T. gondii DNA was found in the organs (heart and brain) of three does. In conclusion, does infected with Toxoplasma gondii in semen at the time of artificial insemination display reproductive disorders in the chronic phase of infection that might be associated with toxoplasmosis.(AU)


Assuntos
Animais , Feminino , Cabras/embriologia , Cabras/parasitologia , Toxoplasma/patogenicidade , Toxoplasmose Animal/complicações , Toxoplasmose Animal/embriologia , Toxoplasmose Animal/fisiopatologia , Doenças Parasitárias em Animais/complicações , Doenças Parasitárias em Animais/patologia , Patologia Veterinária , Infertilidade/veterinária
13.
Rev. bras. ciênc. vet ; 26(1): 21-25, jan.-mar. 2019. ilus, tab
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1491634

Resumo

Objetivou-se descrever os distúrbios reprodutivos associados à infecção experimental por Toxoplasma gondii através da inseminação artificial com sêmen contaminado em quatro cabras no estágio crônico da infecção. As características do trato reprodutor foram avaliadas através de ultrassonografia transretal, visando o diagnóstico gestacional ou de desordens reprodutivas, após a infecção experimental. Ao final do experimento, os animais foram necropsiados e avaliações histopatológicas e PCR foram realizados. Dentre os animais infectados que exibiram mortalidade embrionária, duas apresentaram anestro e duas apresentaram repetição de estro, sendo que destas uma apresentou intervalos entre estros reduzido (sete dias) e outra em intervalo regular (21 dias). Todavia, ambas foram submetidas a monta natural durante os estros naturais subsequentes e não foi confirmada gestação até o final do experimento (90 dias). Duas cabras exibiram alterações nos exames de ultrassonografia, sendo identificadas um cisto ovariano, e uma hidrossalpinge, ambas confirmadas no exame post-mortem. As principais lesões microscópicas nesse grupo foram infiltração neutrofílica dos pulmões, glomerulonefrite intersticial e infiltração neutrofílica do fígado. O DNA de T. gondii foi encontrado nos órgãos (coração e cérebro) de três cabras. Em conclusão, cabras infectadas com sêmen contendo T. gondii no momento da inseminação artificial apresentam distúrbios reprodutivos na fase crônica da infecção que podem estar associados à toxoplasmose.


The aim of this study was to describe the reproductive disorders related to experimental infection by artificial insemination with semen contaminated with Toxoplasma gondii of four goats in the chronic phase of the infection. In the end of the study, the does were submitted to necropsy, and PCR and histopathological evaluations were performed. Among infected does that exhibited embryonic loss, two were in anestrus and two exhibited repeated estrus. One of the latter animals exhibited clinical signs of estrus at seven-day intervals, whereas the other had a 21-day estrous cycle. However, both does were naturally mated on subsequent natural estrous and were not able to get pregnant until the end of the experiment (90 d). Two of the goats exhibited abnormalities in the ultrasound examinations, one of which was an ovarian cyst, while the other was a hydrosalpinx, both of which were confirmed in the post-mortem examination. The main microscopic injuries in this group were neutrophilic infiltration of the lungs, interstitial glomerulonephritis and neutrophilic infiltration of the liver. T. gondii DNA was found in the organs (heart and brain) of three does. In conclusion, does infected with Toxoplasma gondii in semen at the time of artificial insemination display reproductive disorders in the chronic phase of infection that might be associated with toxoplasmosis.


Assuntos
Feminino , Animais , Cabras/embriologia , Cabras/parasitologia , Doenças Parasitárias em Animais/complicações , Doenças Parasitárias em Animais/patologia , Infertilidade/veterinária , Patologia Veterinária , Toxoplasma/patogenicidade , Toxoplasmose Animal/complicações , Toxoplasmose Animal/embriologia , Toxoplasmose Animal/fisiopatologia
14.
Anim. Reprod. ; 15(2): 114-117, Apr.-June.2018. tab, graf
Artigo em Inglês | VETINDEX | ID: vti-16901

Resumo

In some domestic ruminants, contact with females is necessary for normal development of sexual behavior. The aims of this experiment were to determine if rearing bucks isolated from does affects negatively their sexual behavior when adults, and if this negative effect is overcome after four short contacts with females. Sixteen Saanen male kids were maintained during one year in two groups: kids reared in permanent direct contact with four adult goats (FEM;n = 7), and kids that remained isolated from females (ISO; n = 9). When bucks were 12 mo-old, females were removed and both groups were joined in the FEM pen. Nine months later all bucks were individually exposed four times to estrual females for 20 min every 10 days, recording courtship and mounting behaviors. Bucks that were reared with females displayed More courtship behaviors, ejaculations and total mounts(mount attempts, mounts with and without ejaculation; P < 0.0001 for all) than ISO bucks. The combined effect of number of bucks that ejaculated and the time at which they first ejaculated in the first and second tests favored FEM bucks (P < 0.03). It was concluded that the lack of contact with females during the rearing period affects negatively adult bucks sexual performance,an effect that could not be overcome after repeated exposures to estural does.(AU)


Assuntos
Animais , Comportamento Sexual Animal , Cabras/embriologia , Desenvolvimento Sexual , Ruminantes
15.
Anim. Reprod. (Online) ; 15(2): 114-117, Apr.-June.2018. tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461347

Resumo

In some domestic ruminants, contact with females is necessary for normal development of sexual behavior. The aims of this experiment were to determine if rearing bucks isolated from does affects negatively their sexual behavior when adults, and if this negative effect is overcome after four short contacts with females. Sixteen Saanen male kids were maintained during one year in two groups: kids reared in permanent direct contact with four adult goats (FEM;n = 7), and kids that remained isolated from females (ISO; n = 9). When bucks were 12 mo-old, females were removed and both groups were joined in the FEM pen. Nine months later all bucks were individually exposed four times to estrual females for 20 min every 10 days, recording courtship and mounting behaviors. Bucks that were reared with females displayed More courtship behaviors, ejaculations and total mounts(mount attempts, mounts with and without ejaculation; P < 0.0001 for all) than ISO bucks. The combined effect of number of bucks that ejaculated and the time at which they first ejaculated in the first and second tests favored FEM bucks (P < 0.03). It was concluded that the lack of contact with females during the rearing period affects negatively adult bucks’ sexual performance,an effect that could not be overcome after repeated exposures to estural does.


Assuntos
Animais , Cabras/embriologia , Comportamento Sexual Animal , Desenvolvimento Sexual , Ruminantes
16.
Anim. Reprod. (Online) ; 14(2): 413-417, Apr.-June.2017. tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461265

Resumo

For evaluating different methods of long term estrous synchronization followed by fixed time artificial insemination and to select the most efficient method, during the breeding season 160 Persian downy does were equally allocated to groups (n = 20/group). Estrus Was synchronized using controlled internal drug release devices alone (CIDR) or with equine chorionic gonadotropin (CIDR-eCG), intravaginal sponge impregnated with 45 mg fluorgestone acetate alone (Sponge) or with eCG (Sponge-eCG),subcutaneous auricular implant of 2 mg norgestomet alone (Implant) or with eCG (Implant-eCG) or two intramuscular injections of 10 mg prostaglandin F2α10 days apart alone (PGF) or accompany with eCG (PGF-eCG). The dose of eCG was 400 IU and injected intramuscularlly at the end of treatments. Heat detection and fixed time artificial insemination were done 12 and 4 hours after the end of the treatments, respectively. The estrus was detected in 94.7-100% of the does who received different forms of progestagens and estrus response was not affected by eCG. Estrus response in PGF-based groups was significantly lower (P <0.05) than progesterone-based groups. Injection of eCG did not affect fertility, fecundity and prolificacy rates in CIDR, Sponge and Implant groups. It also did not affectparturition rate in implant group. Finally, with respect to estrus response, fertility rate, costs and the simplicity of methods it can be concluded that the use of intravaginal CIDR For 16 days without administration of eCG may be a more efficient method for estrus synchronization in fixed-time artificial insemination programs in Persian downy does.


Assuntos
Feminino , Animais , Cabras/embriologia , Cabras/genética , Sincronização do Estro/genética , Sincronização do Estro/métodos , Inseminação Artificial , Inseminação Artificial/veterinária , Reprodução
17.
Anim. Reprod. (Online) ; 14(4): 1095-1102, Oct.-Dec. 2017. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461305

Resumo

The aims of this study were to investigate the effects of different concentrations of Platelet-derived growth factor-BB (PDGF-BB) on the survival, activation, levels of ROS, and growth of goat preantral follicles enclosed in ovarian tissue. For this, ovarian fragments were cultured for 7 days in Alpha Minimum Essential Medium (α-MEM+ ) with or without PDGF-BB (0, 25, 50 and 100 ng/ml). The results showed that both the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments maintained the percentage of morphologically normal follicles from day 1 to day 7. In addition, the 25 ng/ml PDGF treatment showed a significantly higher percentage of morphologically normal follicles when compared to the other treatments. At day 7, greater (P < 0.05) follicular and oocyte diameters were observed in the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments when compared to the cultured control treatment. On day 7 of culture, all the treatments tested had a significant increase in the percentage of developing follicles when compared to the non-cultured control. However, the percentage of follicle activation, as well as ROS production, were similar (P < 0.05) among the treatments, irrespective of culture time. In conclusion, PDGF-BB improved, in a concentration-dependent manner, follicular survival as well as oocyte and follicular diameter after in vitro culture of goat preantral follicle-enclosed in ovarian tissue fragments.


Assuntos
Animais , Cabras/anatomia & histologia , Cabras/embriologia , Fator de Crescimento Derivado de Plaquetas/administração & dosagem , Fator de Crescimento Derivado de Plaquetas/efeitos adversos , Fertilização in vitro , Folículo Ovariano
18.
Anim. Reprod. ; 14(4): 1095-1102, Oct.-Dec. 2017. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: vti-18213

Resumo

The aims of this study were to investigate the effects of different concentrations of Platelet-derived growth factor-BB (PDGF-BB) on the survival, activation, levels of ROS, and growth of goat preantral follicles enclosed in ovarian tissue. For this, ovarian fragments were cultured for 7 days in Alpha Minimum Essential Medium (α-MEM+ ) with or without PDGF-BB (0, 25, 50 and 100 ng/ml). The results showed that both the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments maintained the percentage of morphologically normal follicles from day 1 to day 7. In addition, the 25 ng/ml PDGF treatment showed a significantly higher percentage of morphologically normal follicles when compared to the other treatments. At day 7, greater (P < 0.05) follicular and oocyte diameters were observed in the 25 ng/ml PDGF and the 50 ng/ml PDGF treatments when compared to the cultured control treatment. On day 7 of culture, all the treatments tested had a significant increase in the percentage of developing follicles when compared to the non-cultured control. However, the percentage of follicle activation, as well as ROS production, were similar (P < 0.05) among the treatments, irrespective of culture time. In conclusion, PDGF-BB improved, in a concentration-dependent manner, follicular survival as well as oocyte and follicular diameter after in vitro culture of goat preantral follicle-enclosed in ovarian tissue fragments.(AU)


Assuntos
Animais , Fator de Crescimento Derivado de Plaquetas/administração & dosagem , Fator de Crescimento Derivado de Plaquetas/efeitos adversos , Cabras/anatomia & histologia , Cabras/embriologia , Fertilização in vitro , Folículo Ovariano
19.
Anim. Reprod. ; 14(2): 413-417, Apr.-June.2017. tab, graf
Artigo em Inglês | VETINDEX | ID: vti-15997

Resumo

For evaluating different methods of long term estrous synchronization followed by fixed time artificial insemination and to select the most efficient method, during the breeding season 160 Persian downy does were equally allocated to groups (n = 20/group). Estrus Was synchronized using controlled internal drug release devices alone (CIDR) or with equine chorionic gonadotropin (CIDR-eCG), intravaginal sponge impregnated with 45 mg fluorgestone acetate alone (Sponge) or with eCG (Sponge-eCG),subcutaneous auricular implant of 2 mg norgestomet alone (Implant) or with eCG (Implant-eCG) or two intramuscular injections of 10 mg prostaglandin F2α10 days apart alone (PGF) or accompany with eCG (PGF-eCG). The dose of eCG was 400 IU and injected intramuscularlly at the end of treatments. Heat detection and fixed time artificial insemination were done 12 and 4 hours after the end of the treatments, respectively. The estrus was detected in 94.7-100% of the does who received different forms of progestagens and estrus response was not affected by eCG. Estrus response in PGF-based groups was significantly lower (P <0.05) than progesterone-based groups. Injection of eCG did not affect fertility, fecundity and prolificacy rates in CIDR, Sponge and Implant groups. It also did not affectparturition rate in implant group. Finally, with respect to estrus response, fertility rate, costs and the simplicity of methods it can be concluded that the use of intravaginal CIDR For 16 days without administration of eCG may be a more efficient method for estrus synchronization in fixed-time artificial insemination programs in Persian downy does.(AU)


Assuntos
Animais , Feminino , Sincronização do Estro/genética , Sincronização do Estro/métodos , Cabras/embriologia , Cabras/genética , Inseminação Artificial , Inseminação Artificial/veterinária , Reprodução
20.
Anim. Reprod. (Online) ; 14(4): 1110-1123, Oct.-Dec. 2017. tab, graf
Artigo em Inglês | VETINDEX | ID: biblio-1461307

Resumo

The birth of cloned goats has been well documented, but the overall goat cloning efficiency by somatic cell nuclear transfer procedures is still low, which may be further intensified in extreme environments. The aim of this study was to produce cloned goats under the conditions of the Brazilian Semi Arid region, in a transgenic program for the expression of human lysozyme in the milk to target childhood diarrhea and malnutrition, comparing the effects of oocyte source, cell type, and embryo reconstruction procedures on in vitro and in vivo embryo survival after cloning by micromanipulation or by handmade cloning. The use of in vitro-matured oocytes resulted in more viable embryos after cloning than in vivo-matured cytoplasts, but no differences in pregnancy rates on day 23 were seen between oocyte sources (77.5 vs. 77.8%, respectively). The presence or absence of the zona pellucida for embryo reconstruction (78.8 vs. 76.0%, respectively) did not affect pregnancy outcome after transfer. However, pregnancy rate on day 23 was higher for embryos chemically activated by a conventional than a modified protocol (88.1 vs. 50.0%), and for embryos reconstructed with mesenchymal stem cells and fetal fibroblasts (100.0 and 93.3%) than with adult fibroblasts (64.7%). Although most pregnancies were lost, the birth of a cloned female was obtained from embryos reconstructed by micromanipulation using non-transgenic control cells and in vitro-matured oocytes with intact zona pellucida, after conventional activation and transfer at the 1-cell stage.


Assuntos
Animais , Cabras/embriologia , Clonagem de Organismos , Clonagem de Organismos/tendências , Técnicas de Maturação in Vitro de Oócitos/veterinária
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