Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 108, Issue 19, Pages 7902-7907Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1019507108
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Funding
- National Institutes of Health [R01-NS050835]
- Damon Runyon Cancer Research Foundation [DRG-1819-04]
- European Molecular Biology Organization [ALTF 851-2005]
- Human Frontier Science Program Organization [LT00805/2006-L]
- Swiss National Science Foundation [PA00P3_124160]
- Swiss National Science Foundation (SNF) [PA00P3_124160] Funding Source: Swiss National Science Foundation (SNF)
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Microinjection of recombinant DNA into zygotic pronuclei has been widely used for producing transgenic mice. However, with this method, the insertion site, integrity, and copy number of the transgene cannot be controlled. Here, we present an integrase-based approach to produce transgenic mice via pronuclear injection, whereby an intact single-copy transgene can be inserted into predetermined chromosomal loci with high efficiency (up to 40%), and faithfully transmitted through generations. We show that neighboring transgenic elements and bacterial DNA within the transgene cause profound silencing and expression variability of the transgenic marker. Removal of these undesirable elements leads to global high-level marker expression from transgenes driven by a ubiquitous promoter. We also obtained faithful marker expression from a tissue-specific promoter. The technique presented here will greatly facilitate murine transgenesis and precise structure/function dissection of mammalian gene function and regulation in vivo.
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