Tetracycline-inducible gene expression in nuclear transfer embryos derived from porcine fetal fibroblasts transformed with retrovirus vectors

Bok Ryul Choi, Bon Chul Koo, Kwang Sung Ahn, Mo Sun Kwon, Jin Hoi Kim, Seong Keun Cho, Kyoung Mi Kim, Jee Hyun Kang, Hosup Shim, Hyuna Lee, Sang Jun Uhm, Hoon Taek Lee, Teoan Kim

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

A critical problem of transgenic livestock production is uncontrollable constitutive expression of the foreign gene, which usually results in serious physiological disturbances in transgenic animals. One of the best solutions for this problem may be use of controllable gene expression system. In this study, using retrovirus vectors designed to express the enhanced green fluorescent protein (EGFP) gene under the control of the tetracycline-inducible promoter, we examined whether the expression of the transgene could be controllable in fibroblast cells and nuclear transfer (NT) embryos of porcine. Transformed fibroblast cells were cultured in medium supplemented with or without doxycycline (a tetracycline analog) for 48 hr, and the induction efficiency was measured by comparing EGFP gene expression using epifluorescence microscopy and Western and Northern blot analyses. After the addition of doxycycline, EGFP expression increased up to 17-fold. The nuclei of transformed fibroblast cells were transferred into enucleated oocytes. Fluorescence emission data revealed strong EGFP gene expression in embryos cultured with doxycycline, but little or no expression in the absence of the antibiotic. Our results demonstrate the successful regulation of transgene expression in porcine nuclear transfer embryos, and support the application of an inducible expression system in transgenic pig production to solve the inherent problems of side-effects due to constitutive expression of the transgene.

Original languageEnglish
Pages (from-to)1221-1229
Number of pages9
JournalMolecular Reproduction and Development
Volume73
Issue number10
DOIs
Publication statusPublished - 2006 Oct
Externally publishedYes

Keywords

  • Doxycycline
  • Enhanced green fluorescent protein (EGFP)
  • Nuclear transfer (NT) porcine embryo
  • Retrovirus vector
  • Tetracycline-inducible expression

ASJC Scopus subject areas

  • Genetics
  • Developmental Biology
  • Cell Biology

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  • Cite this

    Choi, B. R., Koo, B. C., Ahn, K. S., Kwon, M. S., Kim, J. H., Cho, S. K., Kim, K. M., Kang, J. H., Shim, H., Lee, H., Uhm, S. J., Lee, H. T., & Kim, T. (2006). Tetracycline-inducible gene expression in nuclear transfer embryos derived from porcine fetal fibroblasts transformed with retrovirus vectors. Molecular Reproduction and Development, 73(10), 1221-1229. https://doi.org/10.1002/mrd.20543