Drosophila transgenesis

Question 1

How can cloned DNA be introduced into flies?

Answer 1

By germ line transformation using a modified transposable element

Question 2

How can you identify transgenic flies?

Answer 2

By using a selectable marker, e.g. flies mutant for white will have red eyes if they carry the P element

Question 3

Why can’t the helper P element insert into the genome?

Answer 3

Because it lacks an inverted repeat

Question 4

The helper P element cannot insert itself into the genome so why is it needed?

Answer 4

To produce a transposase for the P element plasmid to insert into the host genomic DNA

Question 5

Once the P element as inserted into the host genomic DNA, what happens to the plasmids?

Answer 5

They degrade

Question 6

Is the insertion of a P element stable or unstable?

Answer 6

Stable

Question 7

What happens if the insertion of a P element occurs in the germ cells?

Answer 7

It is transmitted onto the next generation and both their soma and germline will be transgenic

Question 8

Give five uses of transgenic flies in research

Answer 8

Rescue a mutant phenotype
Identify or characterise sequences controlling a gene’s expression
Express a gene and protein of choice
Test the structure or function relation of a protein
Interfere with normal endogenous protein function

Question 9

What is meant by rescuing a mutant phenotype?

Answer 9

Can a cloned gene restore a wildtype phenotype?
E.g. eyeless
Confirms the identity of a cloned gene

Question 10

Why would you want to use transgenic flies to look at the expression of a gene or protein of choice?

Answer 10

To test its effect

See if its expression is necessary or sufficient

Question 11

How would you use transgenic flies to test the structure or function relation of a protein?

Answer 11

For example mutagenise specific amino acids and test the effect in vivo

Question 12

What would you be able to find out about a protein by interfering with the normal endogenous protein function using transgenic flies?

Answer 12

Whether it is dominant-negative or dominant-positive

Question 13

What did the misexpression of eyeless in a transgenic fly show?

Answer 13

That this single gene can trigger the developmental programme for the eye

Question 14

Give three ways in which you can control transgene expression

Answer 14

Drive expression with an inducible heat-shock promoter
Use a characterised drosophila promoter or enhancer to drive expression at a defined time or place
Bipartite system (GAL4/UAS)

Question 15

How would you activate a transgene with a temporal control?

Answer 15

Give 30-60 degrees centigrade heat shock

Question 16

What cells will express a transgene if there is no spatial control?

Answer 16

All cells

Question 17

Give an example of how you can use a characterised drosophila promoter or enhancer to drive expression of a transgene at a define time or place

Answer 17

For example, use the enhancer from the sevenless gene to drive expression in the developing eye

Question 18

What two types of recombination can different arrangements of FRTs produce?

Answer 18

Intermolecular and intramolecular recombination

Question 19

The ‘FLP out’ control of gene expression creates clonal patches of cells expressing gene X, give two ways how can this be used

Answer 19

As a lineage marker, e.g. in brain development

To test the gene action in an unnatural context