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J Vet Sci 2018; 19(3): 434-445  https://doi.org/10.4142/jvs.2018.19.3.434
Production of transgenic pigs using a pGFAP-CreERT2/EGFP LoxP inducible system for central nervous system disease models
Seon-Ung Hwang1,2,†, Kiyoung Eun3,†, Junchul David Yoon1,2, Hyunggee Kim3,*, Sang-Hwan Hyun1,2,*
1Laboratory of Veterinary Embryology and Biotechnology, Veterinary Medical Center and College of Veterinary Medicine, and 2Institute of Stem Cell & Regenerative Medicine, Chungbuk National University, Cheongju 28644, Korea
3Department of Biotechnology, School of Life Sciences and Biotechnology, Korea University, Seoul 02841, Korea
Correspondence to: Tel: +82-43-261-3393; Fax: +82-43-267-3150; E-mails: hg-kim@korea.ac.kr (H Kim), shhyun@cbu.ac.kr (SH Hyun)
The first two authors contributed equally to this work.
Received: July 24, 2017; Revised: October 1, 2017; Accepted: November 24, 2017; Published online: May 31, 2018.
Abstract
Transgenic (TG) pigs are important in biomedical research and are used in disease modeling, pharmaceutical toxicity testing, and regenerative medicine. In this study, we constructed two vector systems by using the promoter of the pig glial fibrillary acidic protein (pGFAP) gene, which is an astrocyte cell marker. We established donor TG fibroblasts with pGFAP-CreERT2/LCMV-EGFPLoxP and evaluated the effect of the transgenes on TG-somatic cell nuclear transfer (SCNT) embryo development. Cleavage rates were not significantly different between control and transgene-donor groups. Embryo transfer was performed thrice just before ovulation of the surrogate sows. One sow delivered 5 TG piglets at 115 days after pregnancy. Polymerase chain reaction (PCR) analysis with genomic DNA isolated from skin tissues of TG pigs revealed that all 5 TG pigs had the transgenes. EGFP expression in all organs tested was confirmed by immunofluorescence staining and PCR. Real-time PCR analysis showed that pGFAP promoter-driven Cre fused to the mutated human ligand-binding domain of the estrogen receptor (CreERT2) mRNA was highly expressed in the cerebrum. Semi-nested PCR analysis revealed that CreERT2-mediated recombination was induced in cerebrum and cerebellum but not in skin. Thus, we successfully generated a TG pig with a 4-hydroxytamoxifen (TM)-inducible pGFAP-CreERT2/EGFPLoxP recombination system via SCNT.
Keywords: genetically modified animals, glial fibrillary acidic protein, nuclear transfer techniques, swine


© 2018 The Korean Society of Veterinary Science.

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