Strong and ubiquitous expression of transgenes targeted into the beta-actin locus by Cre/lox cassette replacement.
Conventional approaches to produce transgenic mice recurrently yield unpredictable patterns and levels of transgene expression, a situation calling for the development of new techniques to overcome these drawbacks in the context of overexpression studies. Here we present an efficient method for rapid and reproducible transgenesis using the recombinase mediated cassette exchange (RMCE) (Bouhassira et al.: Blood 90:3332-3344, 1997) procedure. A lox511-EGFP-TK/neo-loxP cassette was placed under the control of the endogenous mouse beta-actin promoter. Heterozygous mice revealed strong and ubiquitous EGFP expression throughout embryogenesis and adulthood. Reproducibly, the same expression pattern was obtained with RMCE when it was used to replace the EGFP-harboring cassette by ECFP or placental alkaline phosphatase (PLAP) reporter genes (DePrimo et al.: Transgenic Res 5:459-466, 1996). Furthermore, the RMCE procedure proved efficient as well in embryonic stem (ES) cells as directly in zygotes. Our results demonstrate ubiquitous expression of floxed transgenes in the endogenous beta-actin locus and they support the general use of the beta-actin locus for targeted transgenesis.
Shmerling
Doron
D
Novartis Institute of Biomedical Research, Basel, Switzerland.
Danzer
Claus-Peter
CP
Mao
Xiaohong
X
Boisclair
Julie
J
Haffner
Michel
M
Lemaistre
Marianne
M
Schuler
Valerie
V
Kaeslin
Edgar
E
Korn
Reinhard
R
Bürki
Kurt
K
Ledermann
Birgit
B
Kinzel
Bernd
B
Müller
Matthias
M
D000199:Actins / Q000235:genetics*; D000818:Animals; D015870:Gene Expression; D018390:Gene Targeting / Q000379:methods*; D005822:Genetic Vectors; D007150:Immunohistochemistry; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic / Q000235:genetics*; D009865:Oocytes; D010957:Plasmids; D045522:Recombinases / Q000235:genetics / Q000378:metabolism; D013234:Stem Cells; D014162:Transfection; D019076:Transgenes / Q000502:physiology*
D000199:Actins; D045522:Recombinases