Transgenic/Knock-out Mouse Shared Resource

点击翻译按钮获取中文摘要

The primary purpose of the Transgenic/Knock-out Mouse Shared Resource is to provide an efficient and economical means for producing genetically modified mice for members of the Massey Cancer Center. Genetically modified mice (including transgenic mice, in which customized genes are introduced into the genome, or "knock-out/knock-in" mice, in which endogenous genes within the mouse genome are specifically inactivated or modified) have become an indispensable tool in cancer research. The utility of such models ranges from studies of the basic biology of tumorigenesis and progression to the creation of genetically accurate tumor models for the evaluation of novel therapeutic approaches. The Transgenic/Knock-out Mouse Shared Resource offers the following services: Transgenic mouse production: The core generates at least three founder mice for each transgene. Knock-out/knock-in mouse production: The core can perform all or a subset of the following procedures: ES cell electroporation; ES cell culture under appropriate selection conditions; expansion of drug-resistant clones; PCR/Southern blot screening of clones for the correct recombination event; reelectroporation with a ere expression vector to delete floxed selection markers and subsequent rescreening of clones; injection of targeted ES cell clones into blastocysts for chimeric mouse production. For projects in which the core has performed the ES cell targeting, at least two independent ES cell clones are generally injected into blastocysts, and a minimum of five chimeras are generated. Mouse line rederivation: Lines of pathogen-infected mice are rederived by embryo transfer to generate specific pathogen-free mice. Sperm cryopreservation: Lines of mice are cryopreserved by collection, freezing, and long-term liquid nitrogen storage of sperm. When needed, re-establishment of the line can be accomplished by thawing sperm and performing in vitro fertilization of eggs isolated from WT females. Tail DNA genotyping: The core isolates genomic DNA from tail biopsies (or other tissue) provided by the investigator and performs PCR genotyping. Consulting services: The core provides consultation on many aspects of transgenic/knock-out mouse technology, including experimental design, vector design, budgeting for mouse colony maintenance, and preparation of applications to the IACUC for the use of laboratory animals in research. In addition, the core staff provides instruction on overall colony management, including breeding strategies, weaning, ear punching, and tail DNA isolation.