The Cre/loxP system is a strategy for controlling temporal and/or spatial gene expression
through genome alteration in mice. As successful Cre/loxP genome alteration depends on
Cre-driver mice, Cre-reporter mice are essential for validation of Cre gene expression
in vivo. In most Cre-reporter mouse strains, although the presence of
reporter product indicates the expression of Cre recombinase, it has remained unclear
whether a lack of reporter signal indicates either no Cre recombinase expression or
insufficient reporter gene promoter activity. We produced a novel ROSA26 knock-in
Cre-reporter C57BL/6N strain exhibiting green emission before and red after Cre-mediated
recombination, designated as strain R26GRR. Ubiquitous green fluorescence and no red
fluorescence were observed in R26GRR mice. To investigate the activation of tdsRed,
EGFP-excised R26GRR, R26RR, mice were produced through the crossing of
C57BL/6N mice with R26GRR/Ayu1-Cre F1 mice. R26RR mice showed extraordinarily
strong red fluorescence in almost all tissues examined, suggesting ubiquitous activation
of the second reporter in all tissues after Cre/loxP recombination. Moreover, endothelial
cell lineage and pancreatic islet-specific expression of red fluorescence were detected in
R26GRR/Tie2-Cre F1 mice and R26GRR /Ins1-Cre F1 mice, respectively.
These results indicated that R26GRR mice are a useful novel Cre-reporter mouse strain. In
addition, R26GRR mice with a pure C57BL/6N background represent a valuable source of
green-to-red photoconvertible cells following Cre/loxP recombination for application in
transplantation studies. The R26GRR mouse strain will be available from RIKEN BioResource
Center (http://www.brc.riken.jp/lab/animal/en/).
Somatic cell nuclear transfer (SCNT) is a useful technique for creating pig strains
that model human diseases. However, production of numerous cloned disease model pigs
by SCNT for large-scale experiments is impractical due to its complexity and
inefficiency. In the present study, we aimed to establish an efficient procedure for
proliferating the diabetes model pig carrying the mutant human hepatocyte nuclear
factor-1α gene. A founder diabetes transgenic cloned pig was generated by SCNT and
treated with insulin to allow for normal growth to maturity, at which point
epididymal sperm could be collected for cryopreservation. In vitro
fertilization and intrafallopian insemination using the cryopreserved epididymal
sperm resulted in diabetes model transgenic offspring. These results suggest that
artificial reproductive technology using cryopreserved epididymal sperm could be a
practical option for proliferation of genetically modified disease model pigs.
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