Knockout Mice Flashcards
What is a knockout mouse?
A genetically modified mouse where a gene of interest has been inactivated by replacing or disrupting it with artificial DNA.
Who created the knockout mouse technique and in what year?
Capecchi, Evans, and Smithies in 1989.
Why are knockout mice valuable in research?
They provide clues about a gene’s usual biochemical, development, and behavioural roles and function.
List some conditions that knockout mice are used to model.
- Cancer
- Obesity
- Heart disease
- Arthritis
- Substance abuse
- Anxiety
- Depression
- Parkinson’s disease
What is the first step in creating a knockout mouse?
Isolating the gene to be knocked out from a mouse gene library.
What is added to the new sequence engineered for gene knockout?
A reporter gene, such as neomycin resistance gene and herpes tk+.
What type of cells are harvested to create knockout mice?
Embryonic (pluripotent) stem cells from 4-day mouse blastocysts.
What technique is used to insert artificial DNA into chromosomes?
Homologous recombination.
By design, what happens to cells that do not undergo successful homologous recombination?
either die due to lack of neomycin resistance or produce a toxin due to random insertion
+ve and -ve control
What is the likelihood of successful recombination in gene knockout?
Relatively low; most altered cells will be heterozygous for the knockout.
What type of mice are produced after reinjecting genetically altered cells?
Chimera mice.
What does interbreeding heterozygous offspring produce?
Homozygous knockout mice, carrying no functional copy of the original gene.
What can the activity of the reporter gene in knockout mice indicate?
The existing gene’s normal pattern of activity in specific mouse tissues.
What is RNA interference used for?
To silence gene expression.
What is conditional gene modification?
Knocking out genes in specific tissues or at specific times.
What are humanised knock-in mice?
Mice with human genes inserted into their genome.
What are KO rats known for in comparison to KO mice?
Typically present a more typical adult patient pathology.
Name an advantage of using knockout mice in research.
Ability to model human disease and test new drugs/therapies pre-clinically.
What is a disadvantage of knockout mice?
Characteristics in KO mice may differ from those observed in humans.
What percentage of knockout mice are developmentally lethal?
~15%.
Give an example of an experiment using KO mice.
[cite?] Trp53-/- mice with mammary tumours: Notably absent in the first 6 months before dying of lymphomas.
How can gene function in mammary tissue be studied without complications from other tumors?
Using conditional knockouts with loxP or FRT recombination sites.
What alternative method can be used to study mammary tumor formation?
Transplantation of cleared mammary ductal rudiments with Trp53–/– epithelium.
What is the significance of the knockout technique in studying human conditions?
It allows for the knockout of any gene at specific times, which is invaluable in studying genetic factors.
