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Reversible transgene expression in mice: now you see it, now you don’t (1121.1)
Author(s) -
Matthaei Klaus,
Morton Susan,
Chaston Daniel,
Baillie Brett,
Hill Caryl
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.1121.1
Subject(s) - transgene , lac operon , genetically modified mouse , biology , microbiology and biotechnology , phenotype , green fluorescent protein , gene , gene expression , genetics
Transgenic mice can suffer from the “Position effect” whereby the transgene integration site, per se , influences the observed phenotype. Multiple ‘Founder’ animals must therefore be studied to determine the actual phenotype of each transgene. We describe a method whereby a transgene can be switched off and on, allowing reliable validation of phenotype. We modified the endothelial specific Tie2 promoter with Lac O sequences derived from the E. coli Lac O/ Lac I system and generated transgenic mice expressing defective Connexin(Cx)40 molecules, with EGFP as reporter. Transgene expression was down‐regulated by breeding with mice expressing the inhibitor protein Lac I and up‐regulated by Isopropyl β‐D‐1‐thiogalacto‐pyranoside (IPTG) in the drinking water. EGFP was quantified using immunohistochemistry and Western blotting. Blood pressure was measured by tail cuff plethysmography and telemetry. EGFP expression was exclusively endothelial and correlated with transgene copy number. Defective Cx40 expression increased blood pressure. Breeding with Lac I mice significantly reduced EGFP expression and blood pressure by 70‐100%. EGFP expression could be reinstated by IPTG in some, but not all, founder lines in a time dependent manner. We conclude that the Lac O/ Lac I system allows temporal transgene regulation and can reliably control for artifactual phenotypes arising from the “Position effect”.