2013 Scientific Report

Van Andel Research Institute | Scientific Report
Research Interests
The goal of the VARI vivarium and transgenics core is to develop, provide, and support high-quality mouse modeling services
for the VARI investigators, collaborators, and the greater research community. The vivarium is a state-of-the-art facility that
includes a high-level containment barrier. All procedures are conducted according to the NIH Guide for the Care and Use
of Laboratory Animals. Because we understand the importance of excellence in animal care to producing quality research,
we are committed to the highest quality in animal standards, and the Van Andel Research Institute is an AAALAC-accredited
institution. The staff provides rederivation, surgery, dissection, necropsy, breeding, weaning, tail biopsies, sperm and embryo
cryopreservation, animal data management, and health-status monitoring. Transgenic mouse models are produced on request
for project-specific needs.
Transgenics
Fertilized eggs contain two pronuclei, one that is derived from the egg and contains the maternal genetic material and one
derived from the sperm that contains the paternal genetic material. As development proceeds, these two pronuclei fuse,
the genetic material mixes, and the cell proceeds to divide and develop into an embryo. Transgenic mice are produced by
injecting small quantities of foreign DNA (the transgene) into a pronucleus of a one-cell fertilized egg. DNA microinjected into a
pronucleus randomly integrates into the mouse genome and will theoretically be present in every cell of the resulting organism.
Expression of the transgene is controlled by elements called promoters that are genetically engineered into the transgenic
DNA. Depending on the selection of the promoter, the transgene can be expressed in every cell of the mouse or in specific cell
populations such as neurons, skin cells, or blood cells. Temporal expression of the transgene during development can also
be controlled by genetic engineering. These transgenic mice are excellent models for studying the expression and function of
the transgene in vivo.
Gene targeting
Mouse models are produced using gene-targeting technology, a well-established, powerful method for inserting specific
genetic changes into the mouse genome. The resulting mice can be used to study the effects of these changes in the complex
biological environment of a living organism. The genetic changes can include the introduction of a gene into a specific site in
the genome (gene “knock-in”) or the inactivation of a gene already in the genome (gene “knock-out”). Since these mutations
are introduced into the reproductive cells known as the germline, they can be used to study the developmental aspects of gene
function associated with inherited genetic diseases.
The vivarium and transgenics lab can also produce mouse models in which the gene of interest is inactivated in a target organ
or cell line instead of in the entire animal. These models, known as conditional knock-outs, are particularly useful in studying
genes that, if missing, cause the mouse to die as an embryo.
Our gene-targeting service encompasses three major procedures: DNA electroporation, clone expansion and cryopreservation,
and microinjection. Gene targeting is initiated by mutating the genomic DNA of interest and inserting it into mouse embryonic
stem (ES) cells via electroporation. The mutated gene integrates into the genome and, by a process called homologous
recombination, replaces one of the two wild-type copies of the gene in the ES cells. Clones are identified, isolated, and
cryopreserved, and genomic DNA is extracted from each clone and delivered to the client for analysis. Correctly targeted ES
cell clones are thawed, established into tissue culture, and cryopreserved in liquid nitrogen. Gene-targeting mutations are
introduced by microinjection of the pluripotent ES cell clones into 3.5-day-old mouse embryos (blastocysts). These embryos,
containing a mixture of wild-type and mutant ES cells, develop into mice called chimeras. The offspring of chimeras that inherit
the mutated gene are heterozygotes possessing one copy of the mutated gene. The heterozygous mice are bred together to
produce “knock-out mice” that completely lack the normal gene and have two copies of the mutant gene.
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