In vitro development of goat-sheep and goat-goat zona-free cloned embryos in different culture media

Tali

Reprod Domestic Animals

et al. 2018

Conventional somatic cell nuclear transfer (SCNT) technique of in vitro production of cloned embryos involves use of costly and complicated micromanipulators. Handmade cloning (HMC) technique has been applied as efficient and cost-effective alternative in many livestock species. The aim of the present study was to compare the efficiency of in vitro production and in vitro development of cloned sheep embryos by the two techniques. Cloned embryos were produced by conventional SCNT using micromanipulator apparatus and by HMC technique. Enucleation efficiency and efficiency of fusion with somatic cell (nucleus donor) were compared. Cleavage percentage was observed on day 2 of in vitro culture (IVC), and morula and blastocyst percentages were calculated on day 7 of IVC. Higher enucleation efficiency (96.98 ± 1.01 vs. 93.62 ± 1.03; p > .05) as well as fusion efficiency was obtained with HMC technique than with conventional SCNT (96.26 ± 1.34 vs. 92.63 ± 0.70, p < .05); 181 cloned sheep embryos were produced in vitro by conventional SCNT and 92 by HMC. Cleavage percentage observed on day 2 of in vitro culture was higher in HMC than SCNT (66.92 ± 3.72 vs. 55.97 ± 2.5, respectively, p < .05). Morula percentage obtained was higher in SCNT than HMC (44.12 ± 2.93 vs. 30.43 ± 6.79, respectively, p < .05), whereas blastocyst percentage obtained by HMC was higher (12.46 ± 4.96) than SCNT (5.31 ± 2.25; p > .05). It was inferred that HMC technique provides a cost-effective and efficient method of in vitro production of cloned sheep embryos with a comparatively simpler technique with a possibility of automation. Efficiency of cloned embryo production could be improved further by propagating and standardizing this technique.

Section: Discussionmentioning

confidence: 76%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 97%

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Comparison of efficiency of in vitro cloned sheep embryo production by conventional somatic cell nuclear transfer and handmade cloning technique

Tali

Reprod Domestic Animals

et al. 2018

“…Se ha comparado el uso de RVCL (Por sus siglas en inglés Research Vitro Cleave Medium) vs. G1/G2 en clonación manual interespecies (caprino-ovino), encontrando mejores tasas de producción de mórulas y blastocistos (65% vs. 42%, respectivamente) (Khan et al, 2014).…”

Section: Introductionunclassified

Efecto del uso de medio secuencial humano en la producción de blastocistos de hembra ovis canadensis mexicana por clonación manual

Vázquez-Avendaño

Hernández-Martínez

Hernández-Pichardo

et al. 2017

AZM

. (2017). Efecto del uso de medio secuencial humano en la producción de blastocistos de hembra Ovis canadensis mexicana por clonación manual interespecies. Acta Zoológica Mexicana (n.s.), 33(2), 328-338. RESUMEN.Efecto del uso de medio secuencial humano en la producción de blastocistos de hembra Ovis canadensis mexicana por clonación manual interespecies. Desde el evidente problema del peligro de extinción que enfrenta un número significativo de especies silvestres a nivel mundial, se han aplicado biotecnologías encaminadas a promover su conservación y reproducción. Tal es el caso de la transferencia nuclear de células somáticas (TNCS) interespecies que ha sido aplicada y ha producido exitosamente embriones y crías nacidas vivas en algunas de estas especies. Sin embargo, la TNCS interespecies aún debe lidiar con otros problemas como la baja tasa de producción de embriones y de supervivencia de las crías nacidas (3-6%). Problemas que, entre otros, involucran la alta incidencia de fragmentación embrionaria. Los estudios son ahora dirigidos a resolver estos obstáculos, determinando el efecto de diferentes medios o suplementos utilizados en la producción in vitro de embriones. El objetivo de este estudio fue aplicar una variante de la TNCS interespecies (Clonación Manual-HMC) para la producción de embriones, determinando el efecto del uso de medios secuenciales humanos, en el borrego salvaje mexicano ABSTRACT. Effect of sequential human medium use in the production of blastocysts of female Ovis canadensis mexicana by interspecies handmade cloning. Faced to the notable problem that a significant number of world wild species are endangered, some biotechnologies such as the interspecific somatic cell nuclear transfer (SCNT), have been applied to raise their reproduction and conservation. This technology has been successfully applied by producing embryos and live offspring in some of these species. However, the interspecific SCNT still has to solve problems such as the low embryo production and offspring survival rates (3-6%). These problems, among others, involve a high incidence of embryo fragmentation and in order to solve them, most studies are now aimed to determine the effect of different culture media or supplements used for in vitro embryo production. Therefore, the aim of the present study was to use a variant of the interspecific SCNT technique (Handmade cloning-HMC) for embryo production, determining the effect of human sequential media in the endangered Mexican wild sheep O. c. mexicana. Results indicate that the embryo production increased when using human sequential media, allowing EFECTO DEL USO DE MEDIO SECUENCIAL HUMANO EN LA PRODUCCIÓN DE BLASTOCISTOS DE HEMBRA OVIS CANADENSIS MEXICANA

Veterinary Medicine & Sci

Developmental dynamics of cloned Mexican bighorn sheep embryos using morphological quality standards

Martínez

Pichardo

Avendaño

et al. 2020

The developmental dynamics of cloned Mexican bighorn sheep ( Ovis canadensis mexicana ) embryos were evaluated based on morphological quality standards. Categories determined by standards were correlated with the embryonic development stage, number of nuclei and viability. The results showed no differences in the blastocyst rate between the experimental (cloned Mexican bighorn sheep embryos) and control (parthenogenetic domestic sheep embryos) groups ( p > .05), while type IV fragmentation was higher in clones ( p < .05). The standards allowed for the identification of embryos that divided at least once or fragmented after 24 hr of culture. The highest percentage of morulae appeared at 96 hr, the final stages of development: nonsegmented, blocked, fragmented and blastocysts appeared at 192 hr. Embryonic quality decreased over time, making 96 hr the ideal time point to predict the final morphological quality of embryos. Nuclear staining of the morulae and blastocysts showed that higher embryo quality was associated with a higher percentage of normal and viable blastomeres. The evaluated criteria allowed for descriptions of the dynamics, stage and quality of cloned Mexican bighorn sheep embryos with a high degree of reliability. In addition, developmental anomalies, including fragmentation, multinucleation and blocking, were identified.