2 years ago

2013 Scientific Report

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Bryn Eagleson, B.S.,

Bryn Eagleson, B.S., RLATG Vivarium and Laboratory of Transgenics Ms. Eagleson began her career in laboratory animal services with Litton Bionetics at the National Cancer Institute’s Frederick Cancer Research and Development Center (NCI-FCRDC) in Maryland. She later worked at the Johnson & Johnson Biotechnology Center in San Diego, California. In 1988, she returned to the NCI-FCRDC as manager of the transgenic mouse colony. In 1999, Ms. Eagleson was recruited to VARI as the Vivarium Director and Transgenics Special Program Manager. She has a B.S. degree in psychology from the University of Maryland University College. Ms. Eagleson is a member of the IACUC and has served two terms as its chair. Standing, from left: Kefene, Guikema, Boguslawski, Post, Ramsey, Meringa, Baumann, B. Eagleson, Timmer, Stroben, Vrbis, K. Eagleson, Brady, Ehrke Kneeling, from left: Kempston, Rackham, Brandow, Holzgen Research Technicians Laboratory Animal Technicians Animal Caretaker Staff Audra Guikema, B.S., LVT Tristan Kempston, B.S. Kristen Baumann, B.S. Elissa Boguslawski Susan Budnick, B.S. Lisa Kefene, B.S. Tina Meringa, A.S. Janelle Post, B.S. Lisa Ramsey, A.S., LVT Sylvia Timmer, Vivarium Supervisor Crystal Brady Neil Brandow Kendra Eagleson Crystal Ehrke Katie Holzgen Mat Rackham Brandon Stroben Ashlee Vrbis 19

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. 20

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