VL4 - Mouse and Rats as mammalian model organism Flashcards
The laboratory mouse
- Originated from a mixture of Mus musculus domesticus , Mus musculus musculus and Mus musculus castaneus
- Inbred strain is a strain that has in siblingmated for 20 or more generations and can be traced to a single ancestral pair (developed to prove or disaprove the existence of genetic factors)
Importance of inbreeding
Genetic diffrences among individuals impact various aspects
* diseas susceptibility
* reproductive characteristics
* hormone response
* growth perfomance
* immune capacity
* blood components
Reproducibilty is crucial for valid research (>3R) thats why its important to control
* environmental conditions
* cleanliness
* food compositions
* animal demographics
–> important for reliable experimental outcome
Why mice?
- Tolerates inbreed well (many inbred strains available)
- genomic sequenced and annotated
- biologically very similar tu humans,95% of genes in mouse occure in human. (Many of diseases have the same genetic cause as in humans)
- Reproduction: short generation time (3weeks), breed year round, large litters (3-10 baby), ease of cross-fostering
- adapt to social grouping
- short lifespan (2-3 years)
- Omnivorous
Fields of research using the mouse:
* Cancer
* Development and aging-related
* Immunsystem and blood disorder
* Metabolic diseas
* Nurological and sensory disorder
What is PDX model and the use in research
PDX = Patient derived Xenograph
Model for cancer research:
* Implantation of patients cancer cells into immune compromised mouse host e.g: SCID
* in vivo basic cancer biology studies
(Characterization, Tumorigenesis, Metastasis)
* Preclinical cancer research
(Therpaeutic targets, drug screening, drug evaluation)
Tools to study the mouse model
- Staining and imaging
(antibody staining, FISH, histology.. ) - Phenotype alanysis (Behavior e.g: mazes, Physiology e.g: Calorimetry, temp)
- Databases (MGI: Mouse Genome Informations, IMPC: International Mouse Phenotyping Consortium, JAX Mice, Charles River)
Life Cycle of a mouse
Stem cells and trangenesis
Embryoinic cells: totipotent
Give rise to all cell types of the body + extraembryonic cells ( placenta)
Embryonic stemm cells: pluripotent
Give rise to all cell types of the body
Adult stem cells: multipotent
more limited than pluripotent cells but can give rise to more than one cell type
**Induced pluripotent stem cells (iPS) **
adult mature cell + yamanaka factors
What is chimeras?
chimera refers to an organism that contains cells with differnet genetic compositions. Occus naturally or can be artifically introduced through various methods.
Natural: fraternal twins, merging of tissue from diffrent zygotes.
Genetic engineered: creating chimeric animals by introducing cells from one organism into the developing embryo of another.
Human-Mouse Chimeras:
human cells can be introduced into the developing embryos of mice to create human-mouse chimeras. This is done to study human-specific processes or test the potential of human cells in regenerative medicine.
The knock-in approach
Shared Nobel Prize in Physiology and Medicine 2007 to Mario Capecchi, Sir Martin Evans, Oliver Smithies
The first reports in which homologous recombination in ES cells was used to generate
gene-targeted mice were published in 1989.
Genes altered in vitro can thus be passed on to
future generations.
. ESC (Embryonic stem cells) can be used as vehicle to transmit genetic information in mice.
manipulation in culture of embryonic stem cells that are subsequently incorporated back into the embryo proper for shepherding into the germ line
General strategie for generating transgenic mice
microinjection or electroporation of pronucleus of 1 cell embryos.
This foreign DNA usually only integrates into the genome at a random position after the first or second cell division. This means that the transgenic DNA is not present in all cells of the mouse, making it a non-transgenic animal. The transgenic eggs or sperm from these mice are then used to create the next generation of fully transgenic mice.
–>Gene trageting in mice
1. ES cell culture
ES cells are cultivated from mouse pre-implantation embryos (blastocytes)
2. Construction of target vectors
Vector contains pieces of DNA that are homologues to target gene , as well as inserted DNA which changes the target gene and allows for positive-negative selection.
3. ES cell transfection
The Cellular machinery for homologues recombination allows the targeting vector. Enables target vector to find and recombine with target gene.
4. Proliferation of target ES cell
–>gene targeted mice
5. Injection of ES cells into blastocysts
6. form mosaic
7. Birth and breeding of mosaic mice
1981: First transgenic mice. Insertion of foreign DNA into mouse germ line through microinjection of zygote
From gene targeted ES cells to gene targeted mice
Limitations of a knock-out
- Gene mus be known/sequenced
- As a model for human diseases: one mutation in mouse can fail to reproduce the human dieseas for various resons
-Compensation mechanism
-undetected phenotype - Porssible artefact:
-Prolonged Cre expression can be toxic
-randiom insertion in the gneome > variable Cre expression
-can have unexpected activity in off-target tissue - Embryonic lethality:
-Can limit the analysis of the mouse to developmental stages
-less relevant for human health
-some genes can habe diffrent functions in embryos and adults
The laboratory rat
Rattus norvegicus
- physiology is more like the corresponding to human conditions
- bigger animal: physiology easier to monitor, better for surgery, serial blood draws
- rat is more intelligent, behaviour more complex: important for study of human brain development and diseas, cognition and memory research
- cardiovascular disease models superior, particularly for stroke and hypertension
–> genetic tool box for rat is growing
–> Hypertension models inbread strain (e.g: SHR)
The pathogenesis of hypertension in rats and humans are largely similar.
