Genetically modified mouse

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The genetically modified mouse in which a gene affecting hair growth has been knocked out (left) shown next to a normal lab mouse Knockout Mice5006-300.jpg
The genetically modified mouse in which a gene affecting hair growth has been knocked out (left) shown next to a normal lab mouse

A genetically modified mouse or genetically engineered mouse model (GEMM) [1] is a mouse (Mus musculus) that has had its genome altered through the use of genetic engineering techniques. Genetically modified mice are commonly used for research or as animal models of human diseases and are also used for research on genes. Together with patient-derived xenografts (PDXs), GEMMs are the most common in vivo models in cancer research. Both approaches are considered complementary and may be used to recapitulate different aspects of disease. [2] GEMMs are also of great interest for drug development, as they facilitate target validation and the study of response, resistance, toxicity and pharmacodynamics. [3]

Contents

History

In 1974 Beatrice Mintz and Rudolf Jaenisch created the first genetically modified animal by inserting a DNA virus into an early-stage mouse embryo and showing that the inserted genes were present in every cell. [4] However, the mice did not pass the transgene to their offspring, and the impact and applicability of this experiment were, therefore, limited. In 1981 the laboratories of Frank Ruddle [5] from Yale University, Frank Costantini and Elizabeth Lacy from Oxford, and Ralph L. Brinster and Richard Palmiter in collaboration from the University of Pennsylvania and the University of Washington injected purified DNA into a single-cell mouse embryo utilizing techniques developed by Brinster in the 1960s and 1970s, showing transmission of the genetic material to subsequent generations for the first time. [6] [7] [8] During the 1980s, Palmiter and Brinster developed and led the field of transgenesis, refining methods of germline modification and using these techniques to elucidate the activity and function of genes in a way not possible before their unique approach. [9]

Methods

There are two basic technical approaches to produce genetically modified mice. The first involves pronuclear injection, a technique developed and refined by Ralph L. Brinster in the 1960s and 1970s, into a single cell of the mouse embryo, where it will randomly integrate into the mouse genome. [10] This method creates a transgenic mouse and is used to insert new genetic information into the mouse genome or to over-express endogenous genes. The second approach, pioneered by Oliver Smithies and Mario Capecchi, involves modifying embryonic stem cells with a DNA construct containing DNA sequences homologous to the target gene. Embryonic stem cells that recombine with the genomic DNA are selected for and they are then injected into the mice blastocysts. [11] This method is used to manipulate a single gene, in most cases "knocking out" the target gene, although increasingly more subtle and complex genetic manipulation can occur (e.g. humanisation of a specific protein, or only changing single nucleotides). A humanised mouse can also be created by direct addition of human genes, thereby creating a murine form of human-animal hybrid. For example, genetically modified mice may be born with human leukocyte antigen genes in order to provide a more realistic environment when introducing human white blood cells into them in order to study immune system responses. [12] One such application is the identification of hepatitis C virus (HCV) peptides that bind to HLA, and that can be recognized by the human immune system, thereby potentially being targets for future vaccines against HCV. [13]

Uses

Transgenic mice expressing green fluorescent protein, which glows green under blue light. The central mouse is wild-type. GFP Mice 01.jpg
Transgenic mice expressing green fluorescent protein, which glows green under blue light. The central mouse is wild-type.

Genetically modified mice are used extensively in research as models of human disease. [14] Mice are a useful model for genetic manipulation and research, as their tissues and organs are similar to that of a human and they carry virtually all the same genes that operate in humans. [15] They also have advantages over other mammals, in regards to research, in that they are available in hundreds of genetically homogeneous strains. [15] Also, due to their size, they can be kept and housed in large numbers, reducing the cost of research and experiments. [15] The most common type is the knockout mouse, where the activity of a single (or in some cases multiple) genes are removed. They have been used to study and model obesity, heart disease, diabetes, arthritis, substance abuse, anxiety, aging, temperature and pain reception, and Parkinson disease. [16] [17] Transgenic mice generated to carry cloned oncogenes and knockout mice lacking tumor suppressing genes have provided good models for human cancer. Hundreds of these oncomice have been developed covering a wide range of cancers affecting most organs of the body and they are being refined to become more representative of human cancer. [9] The disease symptoms and potential drugs or treatments can be tested against these mouse models.

A mouse has been genetically engineered to have increased muscle growth and strength by overexpressing the insulin-like growth factor I (IGF-I) in differentiated muscle fibers. [18] [19] Another mouse has had a gene altered that is involved in glucose metabolism and runs faster, lives longer, is more sexually active and eats more without getting fatter than the average mouse (see Metabolic supermice). [20] [21] Another mouse had the TRPM8 receptor blocked or removed in a study involving capsaicin and menthol. [17] With the TRPM8 receptor removed, the mouse was unable to detect small changes in temperature and the pain associated with it. [17]

Great care should be taken when deciding how to use genetically modified mice in research. [22] Even basic issues like choosing the correct "wild-type" control mouse to use for comparison are sometimes overlooked. [23]

See also

Related Research Articles

<span class="mw-page-title-main">Genetically modified organism</span> Organisms whose genetic material has been altered using genetic engineering methods

A genetically modified organism (GMO) is any organism whose genetic material has been altered using genetic engineering techniques. The exact definition of a genetically modified organism and what constitutes genetic engineering varies, with the most common being an organism altered in a way that "does not occur naturally by mating and/or natural recombination". A wide variety of organisms have been genetically modified (GM), including animals, plants, and microorganisms.

<span class="mw-page-title-main">Genetic engineering</span> Manipulation of an organisms genome

Genetic engineering, also called genetic modification or genetic manipulation, is the modification and manipulation of an organism's genes using technology. It is a set of technologies used to change the genetic makeup of cells, including the transfer of genes within and across species boundaries to produce improved or novel organisms. New DNA is obtained by either isolating and copying the genetic material of interest using recombinant DNA methods or by artificially synthesising the DNA. A construct is usually created and used to insert this DNA into the host organism. The first recombinant DNA molecule was made by Paul Berg in 1972 by combining DNA from the monkey virus SV40 with the lambda virus. As well as inserting genes, the process can be used to remove, or "knock out", genes. The new DNA can be inserted randomly, or targeted to a specific part of the genome.

Gene knockouts are a widely used genetic engineering technique that involves the targeted removal or inactivation of a specific gene within an organism's genome. This can be done through a variety of methods, including homologous recombination, CRISPR-Cas9, and TALENs.

<span class="mw-page-title-main">Human genetic enhancement</span> Technologies to genetically improve human bodies

Human genetic enhancement or human genetic engineering refers to human enhancement by means of a genetic modification. This could be done in order to cure diseases, prevent the possibility of getting a particular disease, to improve athlete performance in sporting events, or to change physical appearance, metabolism, and even improve physical capabilities and mental faculties such as memory and intelligence. These genetic enhancements may or may not be done in such a way that the change is heritable.

A transgene is a gene that has been transferred naturally, or by any of a number of genetic engineering techniques, from one organism to another. The introduction of a transgene, in a process known as transgenesis, has the potential to change the phenotype of an organism. Transgene describes a segment of DNA containing a gene sequence that has been isolated from one organism and is introduced into a different organism. This non-native segment of DNA may either retain the ability to produce RNA or protein in the transgenic organism or alter the normal function of the transgenic organism's genetic code. In general, the DNA is incorporated into the organism's germ line. For example, in higher vertebrates this can be accomplished by injecting the foreign DNA into the nucleus of a fertilized ovum. This technique is routinely used to introduce human disease genes or other genes of interest into strains of laboratory mice to study the function or pathology involved with that particular gene.

<span class="mw-page-title-main">Rudolf Jaenisch</span> German biologist

Rudolf Jaenisch is a Professor of Biology at MIT and a founding member of the Whitehead Institute for Biomedical Research. He is a pioneer of transgenic science, in which an animal’s genetic makeup is altered. Jaenisch has focused on creating genetically modified mice to study cancer, epigenetic reprogramming and neurological diseases.

<span class="mw-page-title-main">Ralph L. Brinster</span> American geneticist

Ralph Lawrence Brinster is an American geneticist, National Medal of Science laureate, and Richard King Mellon Professor of Reproductive Physiology at the School of Veterinary Medicine, University of Pennsylvania.

In molecular cloning and biology, a gene knock-in refers to a genetic engineering method that involves the one-for-one substitution of DNA sequence information in a genetic locus or the insertion of sequence information not found within the locus. Typically, this is done in mice since the technology for this process is more refined and there is a high degree of shared sequence complexity between mice and humans. The difference between knock-in technology and traditional transgenic techniques is that a knock-in involves a gene inserted into a specific locus, and is thus a "targeted" insertion. It is the opposite of gene knockout.

<span class="mw-page-title-main">Genetically modified animal</span> Animal that has been genetically modified

Genetically modified animals are animals that have been genetically modified for a variety of purposes including producing drugs, enhancing yields, increasing resistance to disease, etc. The vast majority of genetically modified animals are at the research stage while the number close to entering the market remains small.

<span class="mw-page-title-main">Genetically modified mammal</span>

Genetically modified mammals are mammals that have been genetically engineered. They are an important category of genetically modified organisms. The majority of research involving genetically modified mammals involves mice with attempts to produce knockout animals in other mammalian species limited by the inability to derive and stably culture embryonic stem cells.

<span class="mw-page-title-main">Beatrice Mintz</span> American biologist (1921–2022)

Beatrice Mintz was an American embryologist who contributed to the understanding of genetic modification, cellular differentiation, and cancer, particularly melanoma. Mintz was a pioneer of genetic engineering techniques and was among the first scientists to generate both chimeric and transgenic mammals.

<span class="mw-page-title-main">Floxing</span> Sandwiching of a DNA sequence between two lox P sites

In genetics, floxing refers to the sandwiching of a DNA sequence between two lox P sites. The terms are constructed upon the phrase "flanking/flanked by LoxP". Recombination between LoxP sites is catalysed by Cre recombinase. Floxing a gene allows it to be deleted, translocated or inverted in a process called Cre-Lox recombination. The floxing of genes is essential in the development of scientific model systems as it allows researchers to have spatial and temporal alteration of gene expression. Moreover, animals such as mice can be used as models to study human disease. Therefore, Cre-lox system can be used in mice to manipulate gene expression in order to study human diseases and drug development. For example, using the Cre-lox system, researchers can study oncogenes and tumor suppressor genes and their role in development and progression of cancer in mice models.

A knockout mouse, or knock-out mouse, is a genetically modified mouse in which researchers have inactivated, or "knocked out", an existing gene by replacing it or disrupting it with an artificial piece of DNA. They are important animal models for studying the role of genes which have been sequenced but whose functions have not been determined. By causing a specific gene to be inactive in the mouse, and observing any differences from normal behaviour or physiology, researchers can infer its probable function.

<span class="mw-page-title-main">Genome editing</span> Type of genetic engineering

Genome editing, or genome engineering, or gene editing, is a type of genetic engineering in which DNA is inserted, deleted, modified or replaced in the genome of a living organism. Unlike early genetic engineering techniques that randomly inserts genetic material into a host genome, genome editing targets the insertions to site-specific locations. The basic mechanism involved in genetic manipulations through programmable nucleases is the recognition of target genomic loci and binding of effector DNA-binding domain (DBD), double-strand breaks (DSBs) in target DNA by the restriction endonucleases, and the repair of DSBs through homology-directed recombination (HDR) or non-homologous end joining (NHEJ).

<span class="mw-page-title-main">History of genetic engineering</span>

Genetic engineering is the science of manipulating genetic material of an organism. The first artificial genetic modification accomplished using biotechnology was transgenesis, the process of transferring genes from one organism to another, first accomplished by Herbert Boyer and Stanley Cohen in 1973. It was the result of a series of advancements in techniques that allowed the direct modification of the genome. Important advances included the discovery of restriction enzymes and DNA ligases, the ability to design plasmids and technologies like polymerase chain reaction and sequencing. Transformation of the DNA into a host organism was accomplished with the invention of biolistics, Agrobacterium-mediated recombination and microinjection. The first genetically modified animal was a mouse created in 1974 by Rudolf Jaenisch. In 1976 the technology was commercialised, with the advent of genetically modified bacteria that produced somatostatin, followed by insulin in 1978. In 1983 an antibiotic resistant gene was inserted into tobacco, leading to the first genetically engineered plant. Advances followed that allowed scientists to manipulate and add genes to a variety of different organisms and induce a range of different effects. Plants were first commercialized with virus resistant tobacco released in China in 1992. The first genetically modified food was the Flavr Savr tomato marketed in 1994. By 2010, 29 countries had planted commercialized biotech crops. In 2000 a paper published in Science introduced golden rice, the first food developed with increased nutrient value.

Breast cancer metastatic mouse models are experimental approaches in which mice are genetically manipulated to develop a mammary tumor leading to distant focal lesions of mammary epithelium created by metastasis. Mammary cancers in mice can be caused by genetic mutations that have been identified in human cancer. This means models can be generated based upon molecular lesions consistent with the human disease.

Human germline engineering is the process by which the genome of an individual is edited in such a way that the change is heritable. This is achieved by altering the genes of the germ cells, which then mature into genetically modified eggs and sperm. For safety, ethical, and social reasons, there is broad agreement among the scientific community and the public that germline editing for reproduction is a red line that should not be crossed at this point in time. There are differing public sentiments, however, on whether it may be performed in the future depending on whether the intent would be therapeutic or non-therapeutic.

<span class="mw-page-title-main">Frank Ruddle</span> American cell and developmental biologist (1929–2013)

Francis Hugh Ruddle (1929–2013) was an American cell and developmental biologist who was the Sterling Professor at Yale University. Ruddle was an early visionary of the Human Genome Project and created the first genetically modified mouse. He was a pioneer in both human and mouse genetics.

Richard Palmiter is a cellular biologist. He was born in Poughkeepsie, NY, and later went on to earn a BA in Zoology from Duke University and a PhD in Biological Sciences from Stanford University. He is employed with the University of Washington where he is a professor of biochemistry and genome sciences. His current research involves developing a deeper understanding of Parkinson's disease. His most notable research is a collaboration with Dr. Ralph Brinster where they injected purified DNA into a single-cell mouse embryo, showing transmission of the genetic material to subsequent generations for the first time.

<span class="mw-page-title-main">Mouse Models of Human Cancer database</span>

The laboratory mouse has been instrumental in investigating the genetics of human disease, including cancer, for over 110 years. The laboratory mouse has physiology and genetic characteristics very similar to humans providing powerful models for investigation of the genetic characteristics of disease.

References

  1. ↑ Singh, M.; Murriel, C. L.; Johnson, L. (16 May 2012). "Genetically Engineered Mouse Models: Closing the Gap between Preclinical Data and Trial Outcomes". Cancer Research. 72 (11): 2695–2700. doi: 10.1158/0008-5472.CAN-11-2786 . PMID   22593194.
  2. ↑ Abate-Shen, C.; Pandolfi, P. P. (30 September 2013). "Effective Utilization and Appropriate Selection of Genetically Engineered Mouse Models for Translational Integration of Mouse and Human Trials". Cold Spring Harbor Protocols. 2013 (11): 1006–1011. doi: 10.1101/pdb.top078774 . PMC   4382078 . PMID   24173311.
  3. ↑ Sharpless, Norman E.; DePinho, Ronald A. (September 2006). "The mighty mouse: genetically engineered mouse models in cancer drug development". Nature Reviews Drug Discovery. 5 (9): 741–754. doi:10.1038/nrd2110. ISSN   1474-1784. PMID   16915232. S2CID   7254415.
  4. ↑ Jaenisch, R.; Mintz, B. (1974). "Simian virus 40 DNA sequences in DNA of healthy adult mice derived from preimplantation blastocysts injected with viral DNA". Proc. Natl. Acad. Sci. 71 (4): 1250–1254. Bibcode:1974PNAS...71.1250J. doi: 10.1073/pnas.71.4.1250 . PMC   388203 . PMID   4364530.
  5. ↑ Kucherlapati, Raju; Leinwand, Leslie A. (2013). "Frank Ruddle (1929–2013". American Journal of Human Genetics . 92 (6): 839–840. doi:10.1016/j.ajhg.2013.05.012. PMC   3675234 . PMID   24242788.
  6. ↑ Gordon, J.; Ruddle, F. (1981). "Integration and stable germ line transmission of genes injected into mouse pronuclei". Science. 214 (4526): 1244–6. Bibcode:1981Sci...214.1244G. doi:10.1126/science.6272397. PMID   6272397.
  7. ↑ Costantini, F.; Lacy, E. (1981). "Introduction of a rabbit β-globin gene into the mouse germ line". Nature. 294 (5836): 92–4. Bibcode:1981Natur.294...92C. doi:10.1038/294092a0. PMID   6945481. S2CID   4371351.
  8. ↑ Brinster R, Chen HY, Trumbauer M, Senear AW, Warren R, Palmiter RD (1981). "Somatic expression of herpes thymidine kinase in mice following injection of a fusion gene into eggs". Cell. 27 (1 Pt 2): 223–231. doi:10.1016/0092-8674(81)90376-7. PMC   4883678 . PMID   6276022.
  9. 1 2 Douglas Hanahan; Erwin F. Wagner; Richard D. Palmiter (2007). "The origins of oncomice: a history of the first transgenic mice genetically engineered to develop cancer". Genes Dev. 21 (18): 2258–2270. doi: 10.1101/gad.1583307 . PMID   17875663.
  10. ↑ Gordon, J.W., Scangos, G.A, Plotkin, D.J., Barbosa, J.A. and Ruddle F.H. (1980). "Genetic transformation of mouse embryos by microinjection of purified DNA". Proc. Natl. Acad. Sci. USA. 77 (12): 7380–7384. Bibcode:1980PNAS...77.7380G. doi: 10.1073/pnas.77.12.7380 . PMC   350507 . PMID   6261253.{{cite journal}}: CS1 maint: multiple names: authors list (link)
  11. ↑ Thomas KR, Capecchi MR (1987). "Site-directed mutagenesis by gene targeting in mouse embryo-derived stem cells". Cell. 51 (3): 503–12. doi:10.1016/0092-8674(87)90646-5. PMID   2822260. S2CID   31961262.
  12. ↑ Yong KS, Her Z, Chen Q (August 2018). "Humanized Mice as Unique Tools for Human-Specific Studies". Archivum Immunologiae et Therapiae Experimentalis. 66 (4): 245–266. doi:10.1007/s00005-018-0506-x. PMC   6061174 . PMID   29411049.
  13. ↑ "Mouse strain C57BL/6-Mcph1Tg(HLA-A2.1)1Enge". The Jackson Laboratory. Retrieved 2023-01-06.
  14. ↑ "Background: Cloned and Genetically Modified Animals". Center for Genetics and Society. April 14, 2005. Archived from the original on November 23, 2016. Retrieved July 11, 2010.
  15. 1 2 3 Hofker, Marten H.; Deursen, Jan van (2002). Transgenic Mouse . Totowa, New Jersey: Humana Press. pp.  1. ISBN   0-89603-915-3.
  16. ↑ "Knockout Mice". Nation Human Genome Research Institute. 2009.
  17. 1 2 3 Julius, David. "How peppers and peppermint identified sensory receptors for temperature and pain". iBiology. Retrieved 2020-05-14.
  18. ↑ McPherron, A.; Lawler, A.; Lee, S. (1997). "Regulation of skeletal muscle mass in mice by a new TGF-beta superfamily member". Nature. 387 (6628): 83–90. Bibcode:1997Natur.387...83M. doi:10.1038/387083a0. PMID   9139826. S2CID   4271945.
  19. ↑ Elisabeth R. Barton-Davis; Daria I. Shoturma; Antonio Musaro; Nadia Rosenthal; H. Lee Sweeney (1998). "Viral mediated expression of insulin-like growth factor I blocks the aging-related loss of skeletal muscle function". PNAS. 95 (26): 15603–15607. Bibcode:1998PNAS...9515603B. doi: 10.1073/pnas.95.26.15603 . PMC   28090 . PMID   9861016.
  20. ↑ "Genetically engineered super mouse stuns scientists". AAP. November 3, 2007.
  21. ↑ Hakimi P, Yang J, Casadesus G, Massillon D, Tolentino-Silva F, Nye C, Cabrera M, Hagen D, Utter C, Baghdy Y, Johnson DH, Wilson DL, Kirwan JP, Kalhan SC, Hanson RW (2007). "Overexpression of the cytosolic form of phosphoenolpyruvate carboxykinase (GTP) in skeletal muscle repatterns energy metabolism in the mouse". Journal of Biological Chemistry . 282 (45): 32844–32855. doi: 10.1074/jbc.M706127200 . PMC   4484620 . PMID   17716967.
  22. ↑ Crusio, W.E.; Goldowitz, D.; Holmes, A.; Wolfer, D. (2009). "Standards for the publication of mouse mutant studies". Genes, Brain and Behavior . 8 (1): 1–4. doi: 10.1111/j.1601-183X.2008.00438.x . PMID   18778401. S2CID   205853147.
  23. ↑ Mohammed Bourdi; John S. Davies; Lance R. Pohl (2011). "Mispairing C57BL/6 Substrains of Genetically Engineered Mice and Wild-Type Controls Can Lead to Confounding Results as It Did in Studies of JNK2 in Acetaminophen and Concanavalin A Liver Injury". Chemical Research in Toxicology . 24 (6): 794–796. doi:10.1021/tx200143x. PMC   3157912 . PMID   21557537.
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