Module 4: Genetics Flashcards
Describe the differences between outbred and inbred animals, and in which ways different strains can differ from one another, and how they can suffer due to their genetic constitution.
Outbred animals have diverse genetic backgrounds, leading to genetic variation.
* inbred animals are genetically uniform due to mating of closely related individuals.
* Different strains vary in traits like behavior, disease susceptibility, and physiology.
* Inbred animals may suffer from increased incidence of genetic disorders due to homozygosity for harmful alleles, while outbred animals may exhibit diverse
phenotypes but lower risk of genetic diseases.
Describe appropriate breeding programs for laboratory animals.
Appropriate breeding programs for laboratory animals include:
o using controlled mating to maintain genetic consistency and health
o genetic monitoring: regularly assess genetic diversity & inbreeding levels
o record keeping: detailed records of lineage and breeding outcomes.
o health screening: routine health checks to detect and prevent diseases.
* backcrossing = transferring a gene modification from donor to recipient
* Speed congenic= breeding strategy used to rapidly transfer a genetic trait from donor to recipient strain, while minimizing the amount of transferred donor genetic material
* Inbreeding
Describe how genetically altered animals can be generated and how they can be used for scientific research.
Gene modification techniques:
Transgenic animals: a gene from another species is inserted in the genome of the animal
- knock-out: a functioning gene in an animal is made non-functional (no longer encodes a protein), only works as a homozygous version
- Knock-in: a gene version from a foreign species is inserted exactly in the site of the same gene in the homolouge version of the animal (done to make an animal produce the human version of a gene, e.g., a hormone), only works as a homozygous version
How to make transgenic animals:
Knock-in:
- mating –> female mice is euthanized the next day, and the fertilized eggs are flushed from the salpinx and injected with a DNA construct with the desired gene –> another female mouse is mated with a sterilized male –> she is anesthetized, and the manipulated eggs are placed surgically in her salpinx –> gives birth to genetically manipulated pups
- the desired DNA construct is microinjected into one of the pronucleuses (typically the male) of the fertilized egg. The DNA will be inserted at a random location, where an incidental break happens (can only be used for inserting knock-in genes, not knock-out)
- breeding for a couple of generations are needed, as this technique only produces hetero/hemizygtic animals
Knock-out:
- blostcytes of some very specific inbred strains of animals embryonic stem cells can be harvested and grown in culture
- in culture the whole set of gene modification techniques can be applied: after gene modifications specific cultivations can select only those cells in which a targeted mutation occurred, and these can be injected into another blastocyst, which is then injected into an anesthetized pseudopregnant female (mad by mating a sterilized male) –> gives birth to chimaeras (mice containing both WT and gene modified cells)
- hereafter the breeding begins
NEW TECHNIQUES
- CRISPR-cas9: DNA sequences isolated from bacteria, linked to the nuclease cas9, which can produce a double break in DNA
Describe principles of genetic monitoring of laboratory animals.
Inbred strains does not necessarily mean more unexpected diseased animals, as these genes has been sorted out long time ago during the initial inbreeding. However, alot of inbred strains, are bred to specifically express diseases, and these strains needs to be monitored. For instance:
- the SCID mouse, nude mouse and NOG mouse are all immune deficient, and needs to be maintained in an environment highly protected against infections.
- The NOD mouse (non-obese diabetic) wil develop T1D, and their blood glucose levels needs to be monitored, to take the right measurements when diabetes breaks out.
- The most commonly used C57BL/6 mouse is very aggresive, and it is therefore important to form socially harmonious groups
- Some strains mat produce animals that are so affected by the diaease, that it needs to be euthanized for ethical reasons
Explain international standards for nomenclature of laboratory animals.
The Institute of Laboratory Animal Research under the US National Institutes of Health will issue a short code for anyone, who intends to breed rats and mice.
Inbred vs. outbred:
Inbred
- lines are refered to as strain
- names in strains are written like: ‘strain name’ / ‘breeder code’
- substrains names are written in the same way, but with all the relevant breeders with the historically oldest breeder first, and the present breeder last
- F1 hybrids: ‘name of mother strian ‘name of father strain’ F1 / ‘breeders’, and preferbly using the official abreviations to shorten
Outbred
- lines are refered to as stock
- names in stocks are written like: ‘breeder code’ : ‘stock name’
- substocks names are written in the same way, but with all the relevant breeders with the historically oldest breeder first, and the present breeder last
Gene modified animals:
- ‘background strain’ - ‘gene name’
- gene names are written in specific ways, e.g., usually in italics (unless it’s transgene, then it’s brackets around the name instead), human genes are written in all capital letters, while mouse/rat genes are written with first letter capital, the rest as lowercase
- for human transgene: Tg
- for non-homologous: N
- for homologous (knock-in): H
- then the number of the founding line
- then the founders lab code
- / ‘lab code of breeder’
knockout
- ‘background strain’ . ‘donor of stem cells’ - ‘animal gene’ ^ (tm ‘number of founding line’ ‘founders lab code’) / ‘lab code of breeder’
- tm is for ‘targeted mutation’
- em can be used instead of tm for endonuclease mutation, e.g., CRISPR/cas9
