Transgenic and Gene-Targeted Mouse Shared Resource

Jan M. Van Deursen, Ph.D.
Director

Mouse models have become increasingly important tools used to understand the origins and underlying causes of cancer. Previously known as the Animal Models Shared Resource, the Transgenic and Gene-Targeted Mouse Shared Resource provides the infrastructure necessary to generate gene-targeted and transgenic mice as models for cancer research to all participating Mayo Clinic Cancer Center (MCCC) investigators.

Transgenic Mice

Researchers design projects that probe the functional genomics of specific genes. Transgenic mice afford researchers a model uniquely designed to observe the expression patterns of a gene or its mutant variant. Transgenic mouse models enable in vivo demonstration of the involvement of genes of interest in the initiation, progression, and metastasis of cancers.

Most MCCC members use transgenic mice to determine whether and how the overexpression of certain genes (or mutants thereof) play a role in the development of cancer. Most of the genes that MCCC members study through transgenesis are overexpressed in certain types of human cancers.

Transgenic mice are generated by pronuclear microinjection of foreign DNA fragments into one-cell-stage mouse embryos. The Shared Resource provides pronuclear injection of transgenic DNA, and assists investigators with the design of transgenic DNA constructs.

The Shared Resource has a collection of vectors that can be employed for ubiquitous or tissue-specific expression of transgenes. These vectors are freely available to MCCC members. Once a transgenic construct has been assembled, Shared Resource personnel remove the transgene from plasmid backbone and prepare it for microinjection according to a standardized protocol. The DNA construct can then be injected into fertilized eggs. Once pups are born from these eggs, tail biopsies are collected for preparation of genomic DNA. This DNA will be screened for the presence of transgenic DNA by PCR analysis. Animals containing the desired transgene will be transferred to the investigator for further analysis.

Gene-Targeted Mice

Mice with targeted mutations are produced by first modifying a given endogenous gene locus in embryonic stem cells via homologous recombination. Embryonic stem cells carrying the desired gene mutation are then microinjected into blastocyst stage embryos. Microinjected embryos are transferred into pseudo-pregnant host females that develop, deliver and nurse chimeric offspring. The chimeras are then bred to establish the "knockout" mouse line. Gene-targeted mutations can be introduced either in the entire animal or in specific tissues/cell types. These are the standard types of gene-targeted mutations:

• Knockout mutations (classical and conditional gene inactivations)
• Knockin mutations (subtle gene mutations)
• Hypomorphic mutations (down-regulated gene expression)
• Inducible transgenes

We provide two central services:

Gene Targeting Service

The Shared Resource will generate gene-targeted embryonic stem cell clones, inject these clones into blastocysts and breed the resulting chimeras to obtain germline transmission of the targeted mutation.

Blastocyst Microinjection Service

Gene trapping in mouse embryonic stem cells is a powerful and cost-effective approach to create large numbers of insertional mutations in mice. Gene trap embryonic stem cell lines are available from several resources. Shared Resource personnel inject gene trap embryonic stem cells into blastocysts at the request of a MCCC member. The resulting chimeras are then bred to germline transmission of the desired mutation. On average it takes about one year to generate a gene-targeted mouse.