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Dual reporter transgene driven by 2.3Col1a1 promoter is active in differentiated osteoblastsAIM: As quantitative and spatial analyses of promoter reporter constructs are not easily performed in intact bone, we designed a reporter gene specific to bone, which could be analyzed both visually and quantitatively by using chloramphenicol acetyltransferase (CAT) and a cyan version of green fluorescent protein (GFPcyan), driven by a 2.3-kb fragment of the rat collagen promoter (Col2.3). METHODS: The construct Col2.3CATiresGFPcyan was used for generating transgenic mice. Quantitative measurement of promoter activity was performed by CAT analysis of different tissues derived from transgenic animals; localization was performed by visualized GFP in frozen bone sections. To assess transgene expression during in vitro differentiation, marrow stromal cell and neonatal calvarial osteoblast cultures were analyzed for CAT and GFP activity. RESULTS: In mice, CAT activity was detected in the calvaria, long bone, teeth, and tendon, whereas histology showed that GFP expression was limited to osteoblasts and osteocytes. In cell culture, increased activity of CAT correlated with increased differentiation, and GFP activity was restricted to mineralized nodules. CONCLUSION: The concept of a dual reporter allows a simultaneous visual and quantitative analysis of transgene activity in bone.
Document ID
20040087549
Document Type
Reprint (Version printed in journal)
Authors
Marijanovic, Inga (University of Connecticut Health Center Farmington, CT, United States)
Jiang, Xi
Kronenberg, Mark S.
Stover, Mary Louise
Erceg, Ivana
Lichtler, Alexander C.
Rowe, David W.
Date Acquired
August 21, 2013
Publication Date
August 1, 2003
Publication Information
Publication: Croatian medical journal
Volume: 44
Issue: 4
ISSN: 0353-9504
Subject Category
Life Sciences (General)
Distribution Limits
Public
Copyright
Other
Keywords
Non-NASA Center
NASA Discipline Cell Biology