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Development of bison embryos produced by parthenogenesis, interspecies somatic cell nuclear transfer, and ooplasm transfer

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Title: Development of bison embryos produced by parthenogenesis, interspecies somatic cell nuclear transfer, and ooplasm transfer
Author: Gonzalez Grajales, Leslie Antonio
Department: Department of Biomedical Sciences
Program: Biomedical Sciences
Advisor: King, W. Allan
Abstract: The success of reproductive biotechnologies including in vitro embryo production in bison species has been low although several techniques, such as interspecies somatic cell nuclear transfer (iSCNT) hold promise of efficacy in this species. However, interactions between cattle ooplasmic components and bison nuclear structures influence early development in bison iSCNT embryos. Differences in ATP content, incidence of apoptosis, and gene expression between cattle SCNT and bison iSCNT embryos suggest altered interactions in nuclear and mitochondrial genomes, although no differences in developmental rates were observed. A cattle IVM maturation and IVC system were used to determine the developmental potential of plains bison oocytes as cytoplasts. Maturation and developmental rates were similar between cattle and plains bison. The majority of oocytes in both groups reached metaphase meiosis II at 24 hours post maturation and the 8-16 cell stage on day 4. Although total blastocyst rate was similar between cattle and plains bison, embryos in the latter group consistently reached the blastocyst stage 24 to 48 hours after cattle parthenogenic embryos. Modifications to previously described ooplasm transfer techniques were performed to decrease the number of micromanipulations needed when combined with SCNT. The effects of ooplasm transfer into cattle SCNT and iSCNT plains bison embryos was investigated and demonstrated that supplementation of ooplasm matching the genetic origin of the somatic cell in reconstructed SCNT or iSCNT embryos had no effects on development, ATP content, and gene expression profiles at the 8-16 cell stage. Finally, embryo quality between vitrified-thawed wood bison blastocysts produced in vivo and wood bison iSCNT blastocysts generated in vitro was investigated. The percentage of in vivo-derived embryos showing normal morphology was only 20% of that in vitro. Although in vivo embryos expanded, initiated the hatching process, and had higher total cell number, no differences were found in apoptosis incidence when compared to iSCNT embryos. Delayed development at the blastocyst stage was observed in iSCNT wood bison embryos when compared to in vivo embryos. The present study demostrated developmental alterations in iSCNT bison embryos, no effects on development by ooplasm supplementation, and compromised competence of in vivo-derived wood bison embryos.
Date: 2014-12

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