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Integration, stability and expression of the E. coli phytase transgene in the Cassie line of Yorkshire Enviropig™

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Abstract

The genomic structure and generational stability of the transgene carried by the Cassie (CA) line of the transgenic Enviropig™, a prospective food animal, are reported here. This transgene is composed of the Escherichia coli phytase coding sequence regulated by the mouse parotid secretory protein promoter to direct secretion of phytase in the saliva. In the CA line the transgene integrated in chromosome 4 is present as a concatemer of three copies, two in a head to tail orientation and the third in a reverse orientation 3′ to the other copies with a 6 kbp deletion in the 5′ promoter region. The overall size of the integrated transgene complex is 46 kbp. During integration a 66 kbp segment of the chromosome was deleted, but a BLAST search of the segment from a GenBank clone did not reveal any essential genes. The transgene integration site was stable through 9 generations analyzed. Phytase activity in the saliva was similar among 11 day old hemizygous boars and gilts and remained relatively constant through nine generations of hemizygous pigs. However, as the pigs grew there generally was a gradual decrease in activity that stabilized when pigs reached the finisher phase of growth (4–6 months old). Homozygous pigs exhibited 1.5 fold higher phytase activity (P < 0.0001) than that of hemizygous littermates. Moreover, no differential salivary phytase activity was seen in hemizygotes arising from CA-Yorkshire and CA-Duroc breed outcrosses, suggesting that expression of the transgene is unaffected by genetic background. This data demonstrates that an exogenous phytase gene can be stably transmitted and expressed in the salivary glands of a domestic food animal.

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Acknowledgments

This study was funded by research grants from Ontario Pork Producers’ Marketing Board, the Natural Science and Engineering Research Council of Canada, and the Ontario Ministry of Agriculture and Food.

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Author notes

  1. Mingfu Liu

    Present address: LA Biomedical Research Institute, Harbor-UCLA Medical Center, 1224 W Carson St, Torrance, CA, 90745, USA

  2. Serguei Golovan

    Present address: Department of Animal and Food Science, University of Delaware, 531 South College Ave., 046 Townsend Hall, Newark, DE, 19716-2150, USA

Authors and Affiliations

  1. Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON, N1G 2W1, Canada

    Cecil W. Forsberg, Roy G. Meidinger, Michael Cottrill & John P. Phillips

Authors
  1. Cecil W. Forsberg
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  2. Roy G. Meidinger
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  3. Mingfu Liu
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  4. Michael Cottrill
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  5. Serguei Golovan
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  6. John P. Phillips
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Correspondence to Cecil W. Forsberg.

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Forsberg, C.W., Meidinger, R.G., Liu, M. et al. Integration, stability and expression of the E. coli phytase transgene in the Cassie line of Yorkshire Enviropig™. Transgenic Res 22, 379–389 (2013). https://doi.org/10.1007/s11248-012-9646-7

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  • DOI: https://doi.org/10.1007/s11248-012-9646-7

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