Production of pigs expressing a transgene under the control of a tetracycline-inducible system.
Pigs are anatomically and physiologically closer to humans than other laboratory animals. Transgenic (TG) pigs are widely used as models of human diseases. The aim of this study was to produce pigs expressing a tetracycline (Tet)-inducible transgene. The Tet-on system was first tested in infected donor cells. Porcine fetal fibroblasts were infected with a universal doxycycline-inducible vector containing the target gene enhanced green fluorescent protein (eGFP). At 1 day after treatment with 1 µg/ml doxycycline, the fluorescence intensity of these cells was increased. Somatic cell nuclear transfer (SCNT) was then performed using these donor cells. The Tet-on system was then tested in the generated porcine SCNT-TG embryos. Of 4,951 porcine SCNT-TG embryos generated, 850 were cultured in the presence of 1 µg/ml doxycycline in vitro. All of these embryos expressed eGFP and 15 embryos developed to blastocyst stage. The remaining 4,101 embryos were transferred to thirty three surrogate pigs from which thirty eight cloned TG piglets were obtained. PCR analysis showed that the transgene was inserted into the genome of each of these piglets. Two TG fibroblast cell lines were established from these TG piglets, and these cells were used as donor cells for re-cloning. The re-cloned SCNT embryos expressed the eGFP transgene under the control of doxycycline. These data show that the expression of transgenes in cloned TG pigs can be regulated by the Tet-on/off systems.
Department of Animal Science, Chungbuk National University, Cheongju, Republic of Korea.
Yong-Xun
YX
Jin
College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea.
Yubyeol
Y
Jeon
Department of Animal Science, Chungbuk National University, Cheongju, Republic of Korea.
Sung-Hyun
SH
Lee
School of Medicine, Catholic University of Daegu, Daegu, Republic of Korea.
Mo-Sun
MS
Kwon
School of Medicine, Catholic University of Daegu, Daegu, Republic of Korea.
Teoan
T
Kim
Department of Animal Science, Chungbuk National University, Cheongju, Republic of Korea.
Xiang-Shun
XS
Cui
College of Veterinary Medicine, Chungbuk National University, Cheongju, Republic of Korea.
Sang-Hwan
SH
Hyun
Department of Animal Science, Chungbuk National University, Cheongju, Republic of Korea.
Nam-Hyung
NH
Kim
D000818:Animals; D030801:Animals, Genetically Modified; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004624:Embryo Transfer; D004622:Embryo, Mammalian / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D015870:Gene Expression; D017930:Genes, Reporter; D005818:Genetic Engineering; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D053652:Nuclear Transfer Techniques; D030781:Organisms, Genetically Modified; D034421:Sus scrofa / Q000235:genetics*; D013752:Tetracycline / Q000494:pharmacology*; D015533:Transcriptional Activation / Q000187:drug effects*; D019076:Transgenes
C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D013752:Tetracycline