GENETICS - Transgenetics and Cloning

Question 1

What is the difference between a transgenic and cloned animal?

Answer 1

A transgenic animal is created through the deliberate modification of the genome whereas a cloned animal is created through the transfer or an entire genome from one individual to another

Question 2

What are the four main purposes of transgenic animals?

Answer 2

Models for the study of cellular processes
Models for producing human therapeutics
Enhance food quality and production efficiency
Models for developing disease resistant animals and to understand disease processes

Question 3

What are the two ways in which transgenic animals can be genetically modified?

Answer 3

Transgenic animals can have an additional DNA sequence implemented into their genome or they can have a particular DNA sequence deleted or inactivated within their genome

Question 4

What is the term used to describe transgenic animals with a deleted or inactivated DNA sequence within their genome?

Answer 4

Knockout animals

Question 5

What is a transgene construct?

Answer 5

A transgene construct is a experimentally constructed DNA sequence that can be integrated into the genome of the recipient organism to create a transgenic animal

Question 6

What are the three components of a transgene construct?

Answer 6

Promotor sequence
Gene
Poly A sequence

Question 7

What is the function of the promotor sequence in a transgene construct?

Answer 7

The promotor sequence confers the desired spatial and temporal pattern of transgene expression i.e. if the transgene will be expressed in all cells or just in specific tissues

Question 8

What is the typical source of a gene in a transgene construct?

Answer 8

mRNA

Question 9

What are the three techniques used to create transgenic animals?

Answer 9

Pronuclear DNA microinjection
Embryonic cell transfer
Retroviral vectors

Question 10

Describe briefly the process of pronuclear DNA microinjection

Answer 10

1. Eggs are harvested after ovulation is induced
2. Harvested eggs are fertilised in vitro
3. A fine needle is used to inject transgene containing solution into the embryo (usually at the one cell stage of development)
4. Embryos are implanted into a pseudopregnant foster mother
5. After the litter is born, tissue samples are taken from each individual to be analysed for the presence of the transgene using PCR
6. Animals exhibiting optimal expression of the transgene will be selected for breeding so the transgene will remain stable in future generations

Question 11

What are the three advantages of pronuclear DNA microinjection?

Answer 11

1. Simple technique
2. Capable of germline transmission if the transgene is intergrated into reproductive stem cells
3. Can accommodate large transgene constructs

Question 12

What are the three disadvantages of pronuclear DNA microinjection?

Answer 12

1. Random and inefficient integration of the transgene into the genome
2. Only 10 - 40% of the viable offspring are transgenetic
3. More than one copy of the gene can be integrated into the genome which can lead to unstable integration as having multipe copies in close proximity can increase the risk of rearrangements or deletions

Question 13

Describe briefly the process of embryonic cell transfer

Answer 13

1. The transgene is integrated into embryonic stem cells being grown in culture
2. These embryonic stem cells are then integrated into a blastocyst
3. This blastocyst in implanted into a foster mother
4. The litter will exhibit chimerism as the blastocyst is at a multicellular stage of development, so further breeding is required to generate transgenic animals homozygous for the transgene in all cells

Question 14

What are the four advantages of embryonic cell transfer?

Answer 14

Allows for targeting the transgene to a specific insertion site
Can check the location/expression of the transgene in vitro
Can be used to derive knockout animals
The embryonic stem cells can be stored indefinitely

Question 15

What is the main disadvantage of embryonic cell transfer?

Answer 15

The transfer is carried out when the embryo (blastocyst) is at a muticellular stage of development and therefore the animals are mosaic. Because of this, further breeding is required to generate transgenic animals homozygous for the transgene in all cells

Question 16

Describe briefly how retrovirus vectors can be used to create transgenic animals

Answer 16

The transgene constructs are packaged into retrovirus particles and are transferred into the embryos

Question 17

What is the main advantage of using retrovirus vectors to create transgenic animals?

Answer 17

The use of retrovirus vectors allows for the efficient transfer of a single copy of a transgene at a single chromosome location

Question 18

What is the main disadvantage of using retrovirus vectors to create transgenic animals?

Answer 18

The transfer is carried out when the embryo is at a muticellular stage of development and therefore the animals are mosaic. Because of this, further breeding is required to generate transgenic animals homozygous for the transgene in all cells

Question 19

What are the advantages of transgenetics compared to selective breeding?

Answer 19

Increased specificity
Increased speed
Cross species transfer of DNA is possible

Question 20

What are the disadvantages of transgenetics compared to selective breeding?

Answer 20

Potential risk to animal health
Potential risk to the environment

Question 21

What is the advantage of cloning over transgenetics?

Answer 21

Once a desirable transgenic animal is created, this individual will only survive for one generation and there is no guarantee that the offspring of this individual will have as optimal expression of the transgene. However, cloning results in a genetically identical individual

Question 22

Give three examples of what cloning can be used for

Answer 22

Propogate genetic material from sterile/neutered animals
Animal conservation
Harvesting tissues/stem cells

Question 23

What are the main concerns regarding cloning animals?

Answer 23

Cloning has a less than 5% efficiency
The clones often have short lifespans

Question 24

What is stem cell therapy?

Answer 24

Stem cell therapy is the use of stem cells to alleviate disease in animals and humans

Question 25

What is the difference between embryonic and somatic stem cells?

Answer 25

Embryonic stem cells are pluripotent and thus can give rise to all of the cell types within the body whereas somatic stem cells are multipotent and can develop into more than one cell type however are more limited than pluripotent stem cells

Question 26

What are the four main concerns regarding the use of embryonic stem cells for stem cell therapy?

Answer 26

Ethical and moral concerns due to the destruction of embryos
Inefficiency
Expensive
Risk of transmission of animal viruses

Question 27

Why is there a risk of animal virus transmission through using embryonic stem cells for stem cell therapy?

Answer 27

There is a risk of animal virus transmission through using embryonic stem cells for stem cell therapy as these stem cells are cultured in materials which may contain animal products

Question 28

In veterinary medicine, which disease can stem cell therapy be used to treat in cats and dogs?

Answer 28

Arthritis

Question 29

In veterinary medicine, which two diseases can stem cell therapy be used to treat in horses?

Answer 29

Arthritis
Tendon injuries