Testing transgenic regulatory elements through live mouse imaging

Naama Klopstock*, Carol Levy, Devorah Olam, Eithan Galun, Daniel Goldenberg

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

To overcome positional and methylation effects on transgene expression, we developed a universal cloning cassette for in vivo assessment of regulatory elements using the luciferase reporter gene and the CCCD camera. Monitoring luciferase expression pattern in live mice enables screening of large numbers of transgenic founders quickly and inexpensively. We demonstrate that in the engineered transgenic mice, the chicken β-globin 5′HS4 insulator did not always provide the desirable expression pattern, and the Island Element, responsible for the demethylation of the surrounding DNA region, was not beneficial. Both tested liver-specific and developmentally regulated promoters exhibited the expected expression pattern in most transgenic founders.

Original languageAmerican English
Pages (from-to)3986-3990
Number of pages5
JournalFEBS Letters
Volume581
Issue number21
DOIs
StatePublished - 21 Aug 2007
Externally publishedYes

Bibliographical note

Funding Information:
We thank our colleagues from Goldyne Savad Institute of Gene Therapy: Dr. Hila Giladi for help with cloning, Evelyne Zeira for assistance in animal work, Hagit Yotvat for assistance with the CCCD camera, Mery Clausen for assistance on proof-reading and English editing of the manuscript and Meital Guy for secretarial assistance. We thank scientists that kindly provided us with the regulatory elements for gene expression: Prof. Howard Cedar (The Hebrew University, Jerusalem Israel) – for Island Element; Prof. Gary Felsenfeld (NIH, Bethesda, MD) – for chicken β-globin insulator; Dr. Ken-ichi Yamamura (Kumamoto University, Kumamoto, Japan) – for human SAP promoter; Prof. Charles E. Rogler (Albert Einstein College of Medicine, New York, NY) – for murine MUP promoter. E.G. is supported by the Israeli Ministry of Science, through a grant to the National Gene Therapy Knowledge Center and through EC Grants LSHB-CT-2004-512034 (MOLEDA) and LSHB-CT-2005-018961 (INTHER). E.G. is also supported by grants from the Blum, the Harold Grinspoon, the Horowitz and the Wolfson Foundations. D.G. is supported by Kamea Scientific Foundation of the Israeli Government and by Salzberg Foundation.

Keywords

  • CCCD camera
  • In vivo imaging
  • Insulator
  • Liver
  • Promoter
  • Transgene

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