Transgenic Animals Flashcards
What is the traditional method used for improving livestock and other domesticated animals genetically for desirable traits?
Selective breeding through many generations of selective matings.
What are some traits that can be improved through traditional breeding methods?
Milk yield, wool characteristics, rate of weight gain, and egg-laying frequency.
Why is it difficult to introduce new genetic traits using selective-breeding methods once an effective genetic line has been established?
Because a new breeding line may carry both desirable and undesirable genes, and crossing may result in diminished production levels.
What is transgenesis?
Transgenesis is the process of introducing foreign DNA (transgene) into the genetic composition of an animal, resulting in a transgenic animal.
What are some practical applications of transgenic technology?
Studying gene expression and development, establishing animal model systems for studying human diseases, producing foreign proteins in bird eggs, and producing pharmaceuticals in the mammary gland.
Why is the mammary gland often used for producing pharmaceuticals through transgenic technology?
Milk is a renewable, secreted body fluid that can be collected frequently without harm to the animal, and purification of the protein from milk is relatively straightforward due to the small number of different proteins in milk.
What is the disadvantage of using retroviral vectors for transgenesis?
Retroviral vectors are effective in integrating the transgene into the genome of a recipient cell. However, they can only transfer small pieces of DNA and may lack essential adjacent sequences for regulating gene expression
What is a major drawback of using retroviral vectors for transgenesis?
The genome of the retroviral strain used to create the vector DNA can be integrated into the same nucleus as the transgene, potentially leading to retroviral contamination. Additionally, transgenes introduced on retroviral vectors may be silenced in mouse embryos.
What are lentiviruses and why are they used for transgenesis?
Lentiviruses are a group of retroviruses that can deliver large segments of DNA into the host genome, are stable for relatively long periods, have low immunogenicity, and can infect both dividing and nondividing cells. Lentiviral vectors are used for transgenesis because they are capable of delivering larger DNA fragments and do not silence transgenes in embryos.
What are some applications of transgenic mice in biomedical research?
Transgenic mice have been used to study gene regulation, tumor development, immunological specificity, molecular genetics of development, and many other biological processes. They have also been used as biomedical models for various human genetic diseases and to examine the feasibility of industrial production of human therapeutic drugs by domesticated animals.
What are the three methods for introducing DNA into mice for transgenesis?
The three methods for introducing DNA into mice for transgenesis are:
- Retroviral vectors that infect the cells of an early-stage embryo prior to implantation into a receptive female.
- Microinjection into the male pronucleus of a fertilized egg.
- Introduction of genetically engineered embryonic stem cells into an early-stage developing embryo before implantation into a receptive female.
What are some advantages of lentiviral vectors for transgenesis?
Lentiviral vectors have several advantages for transgenesis, including the ability to deliver larger DNA fragments, stability for long periods, low immunogenicity, and the ability to infect both dividing and nondividing cells. They are also not silenced in embryos and do not carry the risk of retroviral contamination.
What is the preferred method for producing transgenic mice and why?
The preferred method for producing transgenic mice is microinjection of DNA into the male pronucleus of fertilized eggs. This method is preferred because it allows for a higher number of available fertilized eggs by stimulating donor females to superovulate, resulting in a larger number of eggs for injection. Additionally, the injected DNA integrates at random sites within the genome, allowing for the creation of transgenic lines of mice carrying functional transgenes.
How is the superovulation of female mice achieved in the microinjection method?
Superovulation of female mice is achieved by giving them an initial injection of pregnant mare’s serum and another injection of human chorionic gonadotropin about 48 hours later. This stimulation results in the production of approximately 35 eggs per superovulated mouse, as opposed to the normal 5 to 10 eggs.
What happens after the fertilized eggs are collected for microinjection?
After the fertilized eggs are collected from the oviducts of superovulated female mice, microinjection of the DNA construct usually occurs immediately. The male pronucleus, which can be located using a dissecting microscope, is injected with the transgene construct, which is often in a linear form and free of prokaryotic vector DNA sequences.
How are the implanted eggs in the foster mother prepared for implantation in the microinjection method?
The foster mother, which has been made pseudopregnant by being mated to a vasectomized male, receives 25 to 40 implanted eggs microsurgically. Copulation is the only known way to prepare the uterus for implantation in mice, and in this case, none of the eggs from the foster mother are fertilized due to the vasectomized mate. The foster mother then delivers pups from the inoculated eggs about 3 weeks after implantation.
How are transgenic animals identified in the microinjection method?
transgenic animals can be identified by assaying DNA from a small piece of the tail using Southern blot hybridization or polymerase chain reaction (PCR) for the presence of the transgene. A transgenic mouse can also be mated to another mouse to determine if the transgene is in the germ line of the founder animal. Subsequently, progeny can be bred with each other to form pure (homozygous) transgenic lines.
What are pluripotent embryonic stem cells?
Pluripotent embryonic stem cells are cells from the blastocyst stage of a developing embryo that can proliferate in cell culture and retain the capability to differentiate into all other cell types, including germ line cells.
How can embryonic stem cells be genetically engineered without altering their pluripotency?
Embryonic stem cells can be genetically engineered by transfection with a DNA vector that is designed to integrate within a specific chromosomal location. This allows for the integration of a functional transgene at a specific site in the genome without affecting pluripotency.
What is the advantage of using the Engineered Embryonic Stem Cell Method over other methods of genetic engineering?
The Engineered Embryonic Stem Cell Method allows for precise genetic engineering without the randomness of integration that is inherent in DNA microinjection and retroviral vector systems. It also avoids interference with essential developmental or cellular functions, as the transgene is integrated into a non-essential region of the genome.
