Expression of human smooth muscle calponin in transgenic mice revealed with a bacterial artificial chromosome

Joseph M. Miano, Chad M. Kitchen, Jiyuan Chen, Kathleen M. Maltby, Louise A. Kelly, Hartmut Weiler, Ralf Krahe, Linda K. Ashworth, Emilio Garcia

Research output: Contribution to journalArticle

Abstract

Defining regulatory elements governing cell-restricted gene expression can be difficult because cis-elements may reside tens of kilobases away from start site(s) of transcription. Artificial chromosomes, which harbor hundreds of kilobases of genomic DNA, preserve a large sequence landscape containing most, if not all, regulatory elements controlling the expression of a particular gene. Here, we report on the use of a bacterial artificial chromosome (BAC) to begin understanding the in vivo regulation of smooth muscle calponin (SM-Calp). Long and accurate polymerase chain reaction, sequencing, and in silico analyses facilitated the complete sequence annotation of a BAC harboring human SM-Calp (hSM-Calp). RNase protection, in situ hybridization, Western blotting, and immunohistochemistry assays showed the BAC clone faithfully expressed hSM-Calp in both cultured cells and transgenic mice. Moreover, expression of hSM-Calp mirrored that of endogenous mouse SM-Calp suggesting that all cis-regulatory elements governing hSM-Calp expression in vivo were contained within the BAC. These BAC mice represent a new model system in which to systematically assess regulatory elements governing SM-Calp transcription in vivo.

Original languageEnglish (US)
Pages (from-to)H1793-H1803
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume282
Issue number5 51-5
StatePublished - Jun 29 2002
Externally publishedYes

Fingerprint

Bacterial Artificial Chromosomes
Transgenic Mice
Smooth Muscle
Artificial Chromosomes
Transcription Initiation Site
Ribonucleases
Computer Simulation
In Situ Hybridization
Cultured Cells
Clone Cells
Western Blotting
Immunohistochemistry
calponin
Gene Expression
Polymerase Chain Reaction
DNA
Genes

Keywords

  • Development
  • Genome
  • Promoter

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

Cite this

Miano, J. M., Kitchen, C. M., Chen, J., Maltby, K. M., Kelly, L. A., Weiler, H., ... Garcia, E. (2002). Expression of human smooth muscle calponin in transgenic mice revealed with a bacterial artificial chromosome. American Journal of Physiology - Heart and Circulatory Physiology, 282(5 51-5), H1793-H1803.

Expression of human smooth muscle calponin in transgenic mice revealed with a bacterial artificial chromosome. / Miano, Joseph M.; Kitchen, Chad M.; Chen, Jiyuan; Maltby, Kathleen M.; Kelly, Louise A.; Weiler, Hartmut; Krahe, Ralf; Ashworth, Linda K.; Garcia, Emilio.

In: American Journal of Physiology - Heart and Circulatory Physiology, Vol. 282, No. 5 51-5, 29.06.2002, p. H1793-H1803.

Research output: Contribution to journalArticle

Miano, JM, Kitchen, CM, Chen, J, Maltby, KM, Kelly, LA, Weiler, H, Krahe, R, Ashworth, LK & Garcia, E 2002, 'Expression of human smooth muscle calponin in transgenic mice revealed with a bacterial artificial chromosome', American Journal of Physiology - Heart and Circulatory Physiology, vol. 282, no. 5 51-5, pp. H1793-H1803.
Miano, Joseph M. ; Kitchen, Chad M. ; Chen, Jiyuan ; Maltby, Kathleen M. ; Kelly, Louise A. ; Weiler, Hartmut ; Krahe, Ralf ; Ashworth, Linda K. ; Garcia, Emilio. / Expression of human smooth muscle calponin in transgenic mice revealed with a bacterial artificial chromosome. In: American Journal of Physiology - Heart and Circulatory Physiology. 2002 ; Vol. 282, No. 5 51-5. pp. H1793-H1803.
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