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Establishment of a bovine blastocyst-derived cell line collection for the comparative analysis of embryos created in vivo and by in vitro fertilization, somatic cell nuclear transfer, or parthenogenetic activation

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Abstract

Tools and methods for analyzing differences in embryos resulting from somatic cell nuclear transfer (NT) in comparison to those derived from normal fertilization are needed to define better the nature of the nuclear reprogramming that occurs after NT. To this end, a collection of bovine blastocyst-derived cell lines was created. In vitro expanded or hatched blastocysts, used as primary culture tissue, were from NT; in vitro maturation, fertilization, and culture (IVF); or parthenogenetic (P) activation. Also, five in vivo-fertilized and developed blastocysts were collected by uterine flushing on the eighth d postfertilization. Whole blastocysts were physically attached to STO feeder layers to initiate all of the cell lines generated. The majority of the cell lines in the collection are trophectoderm, 38 NT-derived, 6 in vivo-derived, 20 IVF-derived, and 13 P-derived. Trophectoderm identity was ascertained by morphology and, in many cases, interferon-tau production. Several visceral endoderm cell lines and putative parietal endoderm cell lines were also established. At approximately 5% efficiency, epiblast masses from NT and IVF blastocysts survived and were isolated in culture. Two epiblast masses were also isolated from P blastocysts. Spontaneous differentiation from the epiblast outgrowths resulted in the establishment of fibroblast cell lines. The use of the trophectoderm cell lines as a comparative in vitro model of bovine trophectoderm and placental function is discussed in relation to NT reprogramming.

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Acknowledgments

We thank Dr. David Guthrie, Dr. Wesley Garrett, Dr. Le Ann Blomberg, Dr. Harris A. Lewin, Dr. Isabelle Hue, and Dr. John M. Talbot for their helpful editorial and scientific comments on the manuscript. We thank Mr. Paul Graninger for technical assistance in embryo production and tissue culture. We also thank Ms. Olga M. Ocon for technical assistance in the assay of IFN-τ. Any mention of trade name, proprietary product, or vendor does not constitute a guarantee or warranty of the product by the US Department of Agriculture or imply its approval to the exclusion of other products or vendors that also may be suitable.

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Authors and Affiliations

  1. USDA, ARS, ANRI, Biotechnology and Germplasm Laboratory, Beltsville, MD, 20705, USA

    Neil C. Talbot & Anne M. Powell

  2. USDA, ARS, Biometrical Consulting Service, Beltsville, MD, 20705, USA

    Mary Camp

  3. Department of Animal Sciences, University of Florida, Gainesville, FL, 32611-0910, USA

    Alan D. Ealy

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  1. Neil C. Talbot
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Correspondence to Neil C. Talbot.

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J. Denry Sato

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Talbot, N.C., Powell, A.M., Camp, M. et al. Establishment of a bovine blastocyst-derived cell line collection for the comparative analysis of embryos created in vivo and by in vitro fertilization, somatic cell nuclear transfer, or parthenogenetic activation. In Vitro Cell.Dev.Biol.-Animal 43, 59–71 (2007). https://doi.org/10.1007/s11626-007-9013-9

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