Lentiviral vectors and LV-Cre-mediated recombination in
vitro and in vivo. (A) Schematic
representation of the lentiviral vectors used in this study.
(Top) The Cre lentiviral vector (LV-Cre) contains a Cre
expression cassette with an nls. Expression of Cre is driven by the CMV
promoter. (Middle) LV-CIG contains an internal ribosomal
entry site (I) and a GFP cassette downstream of Cre.
(Bottom) LV-Lac carries the lacZ gene
instead of Cre. All vectors contain the Rous sarcoma virus promoter in
the 5′ long-terminal repeat (LTR) (for efficient virus production in
the context of a Tat-free packaging system), and self-inactivating
mutations in the 3′ LTR (brown triangle). To enhance transgene
expression in the target cells, a central ppt of HIV-1
pol (ppt) and a posttranscriptional element (W) from the
woodchuck hepatitis virus were included. (B) Structure
of the Cre-responsive lacZ reporter gene unit of CV-1
cells. Before Cre-mediated recombination, transcription from the
CMV-β-actin promoter (thin black arrow) stops at the polyadenylation
site of the neomycin resistance (neo) cassette. After recombination of
the loxP sites (bold black arrows), this cassette is
excised and the lacZ reporter gene is transcribed. Wavy
line, chromosomal DNA. (C and D) X-gal
stain of the CV-1 cells 72 h after infection with LV-Cre (moi
≈ 2) (C) and of uninfected CV-1 cells
(D).