Expression of Nkx2-5-GFP bacterial artificial chromosome transgenic mice closely resembles endogenous Nkx2-5 gene activity

Mouse Nkx2‐5 gene is essential for early heart development and it is regulated by a complex array of regulatory modules. In order to establish an efficient in vivo system for mapping the Nkx2‐5 genomic locus for regulatory regions, we developed improved homologous recombination technology for use in...

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Bibliographic Details
Published in:Genesis (New York, N.Y. : 2000) N.Y. : 2000), 2003-04, Vol.35 (4), p.220-226
Main Authors: Chi, Xuan, Zhang, Shu-xing, Yu, Wei, DeMayo, Francesco J., Rosenberg, Susan M., Schwartz, Robert J.
Format: Article
Language:English
Subjects:
Animals
bacterial artificial chromosome
Chromosomes, Artificial, Bacterial
Gene Expression Profiling
Genes, Reporter
green-fluorescence protein
Homeobox Protein Nkx-2.5
Homeodomain Proteins - genetics
Homeodomain Proteins - metabolism
Mice
Mice, Transgenic
Nkx2-5
Spleen - embryology
Spleen - metabolism
Stomach - embryology
Stomach - metabolism
Transcription Factors - genetics
Transcription Factors - metabolism
transgenic mice
Xenopus Proteins - genetics
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Summary:Mouse Nkx2‐5 gene is essential for early heart development and it is regulated by a complex array of regulatory modules. In order to establish an efficient in vivo system for mapping the Nkx2‐5 genomic locus for regulatory regions, we developed improved homologous recombination technology for use in Escherichia coli and then knocked an IRES‐hrGFP reporter gene into Nkx2‐5 gene in a 120 kb Nkx2‐5 bacterial artificial chromosome (BAC) clone. We employed the recombination genes redα and redβ under the pBAD promoter, which was specifically induced by the addition of L‐arabinose. Recombination was selected for by our universal targeting cassette which conferred kanamycin resistance in bacterial cells and neomycin resistance in mammalian cells. Transgenic mouse lines generated from this modified BAC clone closely resembled the endogenous Nkx2‐5 expression in the heart, pylorus sphincter, and spleen, but expression was not detected in the tongue. Nkx2‐5 BAC‐GFP expression was copy number‐dependent and locus site‐independent. BAC transgenics using the GFP reporter offers an efficient model system to study gene expression and regulation. genesis 35:220–226, 2003. © 2003 Wiley‐Liss, Inc.
ISSN:1526-954X
1526-968X
DOI:10.1002/gene.10181