Document Detail


Establishment of life-span extended bovine fibroblast cells carrying the characterization of primary cells.
MedLine Citation:
PMID:  15529005     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Although primary bovine embryonic fibroblast (BEF) cells have previously been used as nucleus-donors for nuclear transfer (NT), it has now been proposed to use BEF cells to generate cloned cows that were genetically modified by transgenic or a knock-out system. A major limitation to gene targeting somatic cells, however, is the overall life-span of the cell. In this study, we first examined in vitro life-span of primary BEF cells. Primary BEF cells were found to be replicative senescent at passage 10th-12th, similar to primary murine embryonic fibroblast cells. To overcome this short in vitro life-span, we have optimized culture conditions to extend the life-span and determined growth characteristics of BEF cell lines. Two life-span extended BEF cell lines (designated CGFR -BO-1 and CGFR-BO-2) were shown to grow much faster than their parental primary counterparts. Both cell lines did not display any potential for abnormal growth such as foci formations in either soft-agar or confluent culture condition. In cloning experiments using these cell lines as a nuclear donor, the reconstructed karyoblasts underwent apoptosis, reprogramming and development in the blastocyst stage, at a similar frequency to those observed with parental as well as adult primary fibroblasts. Furthermore, these cell lines targeted with green fluorescence protein (GFP) were successfully transduced, selected and reprogrammed by NT to develop into a blastocyst stage with GFP expression. Our results suggested methods to extend life-span of donor cells with tremendous implications for the genetic engineering of bovine fibroblast cells.
Authors:
Seungkwon You; Minhee Heo; Jai-Hee Moon; Sung-Chan Kim; Sungwook Kwak; Du-Hak Yoon; Dongil Jin; Ki-Chang Hong; Douglas N Foster; Yun-Jaie Choi; Hyunggee Kim
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Molecules and cells     Volume:  18     ISSN:  1016-8478     ISO Abbreviation:  Mol. Cells     Publication Date:  2004 Oct 
Date Detail:
Created Date:  2004-11-05     Completed Date:  2005-04-05     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  9610936     Medline TA:  Mol Cells     Country:  Korea (South)    
Other Details:
Languages:  eng     Pagination:  261-8     Citation Subset:  IM    
Affiliation:
Laboratory of Cell Growth and Function Regulation, Korea University, Seoul 136-701, Korea. bioseung@korea.ac.kr
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MeSH Terms
Descriptor/Qualifier:
Animals
Apoptosis
Blastomeres / cytology
Cattle
Cell Aging*
Cell Culture Techniques
Cell Differentiation
Cell Lineage
Cells, Cultured
Cloning, Organism / methods
Fibroblasts / cytology*
Green Fluorescent Proteins / genetics
Nuclear Transfer Techniques
Transduction, Genetic
Chemical
Reg. No./Substance:
147336-22-9/Green Fluorescent Proteins

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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