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Reversible gene silencing in mice
Mirimus has fused together unparalleled expertise of RNA interference technology and genetic engineering of mouse models to create advanced platforms for rapid and cost-effective production of customized RNAi mouse models with potent, reversible gene silencing.
With our latest multEmiR technology, combination gene silencing is now possible. Using our tandem shRNA design, investigators can interrogate entire metabolic pathways or gene families and look for synergistic effects of combination gene silencing.
Our innovative pipeline eliminates the long lead-time for multiple ES cell targeting modifications, mouse generation, intercrossing, and functional validation. Our mouse models provide enormous predictive power that will shape the development of better tolerated therapies.
Gene suppression by RNAi can mimic drug therapy, enabling target identification and validation in both in vitro and in vivo systems.
Inducible RNAi mice enable toxicity assessment to study novel gene targets.
The power of reversible gene silencing
Activation of the Wnt pathway, often through deregulation of APC, is one of the most frequent events in human cancers. To model this event genetically in animals, we inducibly silenced Apc using RNAi mice harboring TRE-shApc/Rosa26-rtTA2 by treating them with doxycycline. After 4-6 weeks of doxycycline treatment, all mice developed primary leukemia/lymphomas, which were transplanted to secondary recipients. Shown in the video is one animal that presented with lymphoma associated hind limb paralysis at 4 weeks post transplantation. After 3 days following doxycycline withdrawal to allow for Apc expression to return to normal endogenous levels, the mouse regained full mobility. This illustrates the power of reversibility using RNAi. Our shAPC RNAi mice enabled us for the first time, to explore the role of APC suppression and its continued deregulation in tumor maintenance.
(A) RNAi mouse generation begins with identification and validation of potent shRNAs, followed by embryonic stem cell targeting and blastocyst injection for mouse generation.
(B) Embryonic stem cell targeting uses recombinase-mediated cassette exchange to precisely incorporate Tet-inducible shRNAs downstream of the Col1a1 locus.
(C) Mice can be treated with doxcycline (a tetracycline analog) through their food or drinking water to induce GFP-coupled shRNA expression.
(D) Western blot analyses of hepatocytes isolated from shMKK4 RNAi mice treated with dox for 5 days.
(E) Whole tissue analyses of tGFP expression in RNAi mice treated with dox for 5 days. CAG-rtTA3 confers global tGFP-shRNA expression. Tissue-specific expression can be obtained by use of alternative tet-transactivator strains or CAG-LSL-rtTA3 and tissue-specific CRE-driver strains.
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