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Mechanism of prolonged gene expression by Epstein–Barr virus‐based plasmid in porcine cells
Author(s) -
Son Jung Kyu,
Oh Sang Taek,
Cho Sung Kyu,
Yoon Kun Ho,
Lee Suk Kyeong
Publication year - 2006
Publication title -
xenotransplantation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.052
H-Index - 61
eISSN - 1399-3089
pISSN - 0908-665X
DOI - 10.1111/j.1399-3089.2006.00350.x
Subject(s) - transfection , plasmid , microbiology and biotechnology , biology , xenotransplantation , cell culture , gene , virus , virology , genetics , transplantation , medicine , surgery
Background: We previously showed that an Epstein‐Barr virus (EBV)‐based plasmid, pEBVGFP, exerts prolonged gene expression in porcine neonatal pancreatic cell clusters (NPCCs). In this study, the mechanism underlying this was investigated. Methods: GFP expression was analyzed in porcine cells transfected with pEBVGFP by FACS analysis and confocal microscopy. The possible integration of pEBVGFP into the chromosomal DNA was analyzed by Southern blot. Self‐replication of the EBV‐based plasmid in porcine cells was investigated by PCR. The NPCCs were immunostained to characterize cells transfected with pEBVGFP. Results: The EBV based plasmid provided prolonged GFP expression in porcine cells and duct cells were the main cells transfected among NPCCs. Southern blot showed that the transfected pEBVGFP stayed for a long time as an episome rather than integrating into the chromosomal DNA. pEBVGFP isolated from the transfected porcine cells had methylated CpG suggesting that they self‐replicated in those cells. Conclusions: The EBV‐based plasmid may be useful for genetically manipulating porcine cells to enhance their value as xenotransplantation sources.