Outcome 5

Question 1

What is a model organsim

Answer 1

A non-human species studied to understrand a fundamental biological phenomena or provide insights into other organisms

Question 2

Which model organism is useful for studying genetic inheritance

Answer 2

Pea Plant for Mendelian inheritance or fruit fly

Question 3

Which model organism is useful for studying cellular processes

Answer 3

Yeast

Question 4

Which animal model is particularly useful for studying embryology

Answer 4

Zebrafish

Question 5

Give three traits/properties of a good model organism.

Answer 5

Quick generation time, easy to grow.

Similarity in physiology, genetics, anatomy when compared to humans.

Question 6

Name the most commonly used mammals for experimental studies of human disease?

Answer 6

Rodents - house mouse and brown rat

Question 7

What is meant by the 3 Rs? Describe them.

Answer 7

Reduction, refinement, replacement.
Reduction - reduce number of animals needed to conduct experiment with improved experimental design. Share information with other researchers.
Refinement - reduce suffering by using less invasive techniques. Anaesthitising animals when possible. Providing better living conditions and handling.
Replacement - Alternatives to model organisms used when possible such as cell cultures, human volunteers.

Question 8

What are some limitations of using an animal model

Answer 8

Some results obtained on animals are not confirmed in further human studies due to differences in physiology, genetics or anatomy.
Genetic and physiological variations within a species can affect the results of an experiment.

Question 9

What is meant by an inbred strain? What is the advantage of using inbred strains (e.g.
of mice) for experiments?

Answer 9

Laboratory mice have been developed as inbred strains which have highly homogeneous genetic composition to increase the reproducibility of results and the statistical power of experiments.

Question 10

Define the term transgene.

Answer 10

Transgene refers to foreign DNA that is transferred into an animal via DNA recombinant technology and is subsequently transmitted through the germ line so that every cell in the body contains the modified genetic material.

Question 11

Name the three principal methods for the creation of transgenic animals?

Answer 11

DNA microinjection
Embryonic stem cell mediated gene transfer
Retrovirus mediated gene transfer

Question 12

Describe the 1st step in the process for generating a transgenic mouse using DNA microinjection

Answer 12

Make the transgene - using recombinant DNA methods, the desired gene, vector DNA, and promoter and regulatory sequences

Question 13

Describe the 2nd step in the process for generating a transgenic mouse using DNA microinjection

Answer 13

In vitro fertilisation - female mouse is superovulated and eggs are fertilised in vitro

Question 14

Describe the 3rd step in the process for generating a transgenic mouse using DNA microinjection

Answer 14

Transform fertilised eggs - male pronucleus is injected with the recombinant DNA. Once the pronuclei have fused to form a diploid zygote nucleus, the zygote divides to form a 2 cell embryo

Question 15

Describe the 4th step in the process for generating a transgenic mouse using DNA microinjection

Answer 15

Implant embryos in pseudopregnant mother - The embryo is transferred into the oviduct of the recipient and the foster mother is induced to act as a recipient by mating with a vasectomised male

Question 16

Describe the 5th step in the process for generating a transgenic mouse using DNA microinjection

Answer 16

Test offspring for transgene - remove tissue and test for transgene via PCR. Heterozygous offspring are mated with homozygous transgenic strain to produce homozygous transgenic strain.

Question 17

What is meant by the term chimera?

Answer 17

An organism composed of cells from more than one distinct genotytpe

Question 18

What is the first step in the process of embryonic stem cell mediated gene transfer

Answer 18

Embryonic ctem cells are harvested from the inner cell mass of the animals blastocyst

Question 19

What is the 2ndt step in the process of embryonic stem cell mediated gene transfer

Answer 19

Make the transgene - recombinant DNA methods used to build a DNA molecule containing the desired gene, vector DNA, promoter and regulatory sequences

Question 20

What is the 3rd step in the process of embryonic stem cell mediated gene transfer

Answer 20

Transform the embryonic stem cells - mix the cells with the recombinant DNA, and perform electroporation. Select for transformed cells

Question 21

What is the 4th step in the process of embryonic stem cell mediated gene transfer

Answer 21

Inject the transformed cells into the inner cell mass of the organisms blastocyst

Question 22

What is the 5th step in the process of embryonic stem cell mediated gene transfer

Answer 22

Implant the embryos into the pseudopregnant mothers oviduct, induce foster mother to act as recipient by mating with vasectomised male

Question 23

What is the 6th step in the process of embryonic stem cell mediated gene transfer

Answer 23

Test offspring for transgene, the offspring are chimeric

Question 24

What is the 7th step in the process of embryonic stem cell mediated gene transfer

Answer 24

Establish a transgenic strain - mate chimera with an albino mouse. Offspring tested to confirm presence of transgene via PCR. If transgene is present then the offspring are heterozygous. MAte heterozygous transgene containing offspring together to establish homozygous transgenic strain

Question 25

What is the major advantage of using retroviruses to mediate gene transfer?

Answer 25

The retrovirus acts as a vector, retroviruses are well evolved to infect host cells

Question 26

Describe the process of retrovirus mediated gene transfer

Answer 26

The retrovirus is used as a vector for the gene of interest, the retrovirus infects the germline (random insertion). Offspring are often chimeric because not all cells are infected. Germ line chimeras are then inbred for 10- 20 generations until homozygous transgenic animals are produced where the transgene is present in every cell

Question 27

Define random gene insertion

Answer 27

The gene is inserted into the animal at a random gene locus and a random number of times (Retrovirus mediated)(microinjected)

Question 28

Define targeted gene insertion

Answer 28

The gene is transfered into the genome at a specific locus (ES mediated) In the vector, the gene of interest (and promoters/enhancers/selection marker) must be flanked by sequences that are homologous to the targeted gene/genome sequence

Question 29

What are knockout mice

Answer 29

When targeted gene is replaced with a nonfunctional gene, no/less protein expression is caused and loss of function. Used to study the function of genes and their role in disease

Question 30

What are knock-in mice

Answer 30

Knock-in mice are produced by inserting a transgene into an exact location where it is over expressed.

Question 31

Outline how animal models can be used to investigate genetic disease with regards to disease mechanism

Answer 31

As animals often do not suffer from the same diseases as humans, the generation of transgenic animals (usually transgenic mice) is often necessary to investigate genetic disease. This is useful as model animals can have genomes similar enough to humans to use them for research into a genetic disease which is linked to a gene mutation (sickle cell anemia, cancers) The transgenic animal can be used to study the mechanism of the disease as well as how the disease progresses. Expression of the gene can be studied to learn in what cells and when it is expressed. The effect of drugs or new trreatments can be studied using the model animal, with a view to confirming finding with later human clinical trials.

Question 32

Outline how animal models can be used to investigate genetic disease with regards to analysing gene expression

Answer 32

The function of many human genes are still poorly understood. By using knockout mice, the effects of selectively removing a gene (and therefore stopping corresponding protein production) can be observed. Humans and mice share many genes and the results may therefore be relevant to humans. Knockout mice have been used to study different types of cancer and Parkinsons disease. The p53 gene supresses tumour development by arresting cell development. By selectively removing this gene in knockout mice, the role that mutations to the p53 gene in humans (such as increased risk of bone cancer) can be better understood