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Establishment of a porcine Oct-4 promoter-driven EGFP reporter system for monitoring pluripotency of porcine stem cells
- Source :
- Cellular reprogramming. 13(2)
- Publication Year :
- 2011
-
Abstract
- Porcine pluripotent cells with the capacity to generate germ line chimeras have not been developed yet. The transcription factor Oct-4 is an important marker of undifferentiating status and a central regulator of pluripotency in cells. Establishment of an Oct-4 promoter-based reporter system, such as that used in mice, will be a useful tool for monitoring the differentiating statuses of porcine cells both in vivo and in vitro. In the present study, we constructed a vector, pOGN2, in which enhanced green fluorescent protein (EGFP) was driven by the porcine Oct-4 promoter. In pigs containing this vector, EGFP was expected to be specifically expressed in pluripotent cells. We delivered the vectors into porcine fetal fibroblasts (PEFs) using liposomes. After transfected PEFs were selected with G418, we established eight cell lines containing the pOGN2 vector. When transgenic cells were used as donor nuclei to make somatic cell nuclear transfer (SCNT) embryos, SCNT embryos derived from four transgenic cell lines expressed green fluorescence. When PEFs with pOGN2 vectors were infected with retroviral vectors encoding the four transcription factors (Oct-4, Sox2, Klf4, and c-Myc), EGFP-expressing iPS cell colonies were observed at day 20. This work lays a foundation that can be used to generate a pig strain with an Oct4-EGFP reporter system, which would be greatly helpful in studying the differentiating and reprogramming mechanisms of pig embryos.
- Subjects :
- Pluripotent Stem Cells
Nuclear Transfer Techniques
Swine
Cellular differentiation
Transgene
Genetic Vectors
Green Fluorescent Proteins
Biology
Oct-4
Green fluorescent protein
Kruppel-Like Factor 4
Mice
Genes, Reporter
Transduction, Genetic
Animals
Promoter Regions, Genetic
Cells, Cultured
Cell Nucleus
Cell Differentiation
Cell Biology
Transfection
Cell Dedifferentiation
Fibroblasts
Molecular biology
Retroviridae
Cell culture
Somatic cell nuclear transfer
Stem cell
Octamer Transcription Factor-3
Developmental Biology
Biotechnology
Transcription Factors
Subjects
Details
- ISSN :
- 21524998
- Volume :
- 13
- Issue :
- 2
- Database :
- OpenAIRE
- Journal :
- Cellular reprogramming
- Accession number :
- edsair.doi.dedup.....13189e599cd8d1f7d41305aa2d393897