Drosophila Embryogenesis
INTRODUCTION
The fruit fly, Drosophila melanogaster, is one of the most commonly used model organisms in
biology. They are used to study everything from molecular genetics to behavior and sexual
dimorphism. Drosophila are easily manipulated by researchers and many mutants exist that
are analogous to human disorders. In addition, Drosophila genes are easily mapped using
their polytene chromosomes, which form through many rounds of duplication but no cell
division (endoreplication). The many copies of chromosomes group together with like parts and
produce banding patterns that give clues to additions or deletions of the genetic
material. Chromosomal puffs, or sections of diffuse, uncoiled DNA, are regions with active RNA
transcription.
Drosophila have three body regions: head, thorax, and abdomen, a trait shared by all insects.
All appendages used for movement originate on the thorax. One pair of wings is located
anterior to the halteres, a modified wing structure used for balance in flight.
Drosophila belongs to the holometabolous group of insects that undergo complete
metamorphosis. Most organs present in the larval stage do not persist into the adult life
stages. Drosophila undergo three larval stages referred to as instars. The eggs hatch in the food
medium and each instar gains progressively more mass as it consumes the food medium. The
third instar forms a pupa, and the adult fly ecloses from this stage. Although most larval tissues
are broken down during the metamorphosis process in the pupa, the larval imaginal
discs persist. The discs are bundles of stem cells that develop into the adult organs during
metamorphosis.
There are several features you can use to determine
whether the fly you are looking at is male or
female. Adult female flies are larger than males and
have several small black stripes on their
abdomen. Their abdomens are more oblong and
taper into a point at the end. Male flies have a black
patch on their abdomen and are more cylindrical in
shape. Male genitalia will look like a round, red disk
at the base of the abdomen whereas female genitalia
will be less obvious.
We will be using the Drosophila model to observe
early larval development and mating behavior. You
will also have a chance to identify various types
of Drosophila mutants and look at prepared slides of the polytene chromosomes.
EXERCISE 1: TA Slideshow
Observe the slide show that your TA will present to familiarize yourself with
wildtype Drosophila anatomy and the differences exhibited by the classical mutants.
EXERCISE 2: Drosophila Mating Behavior
Drosophila is only one of several organisms in which behavior can be directly linked
to genetics. Drosophila males have a very well documented courtship dance that involves
distinct steps. Mutating certain genes can cause males to omit some or all of the steps in the
courtship dance.
Drosophila courtship:
A. Male approaches female anteriorly from angle
B. Male taps female posteriorly with his sex comb
C. Male vibrates wing for female
D. Male licks female’s genitals
E. Male attempts to mount
F. Male copulates with female
Upon copulation the male transfers a sex peptide in addition to sperm, which prevents the
female from mating for a short time.
Vials of male and female virgin flies have been provided for you. Tap out the virgin female flies
into the virgin male fly vial as described in Exercise 2. Place the fly vial on its side and observe
the courtship behavior. Look at adult flies from Exercise 2 as they may be engaging in mating
behavior as well.
When you are finished with the mating behavior exercise, tap the flies into the “fly morgue”
flask on the TA bench and place the vials into the “Used Vials” bin.
QUESTIONS:
1. How long did it take for the first coupling to occur?
2. Once copulation began, how long did it last?
3. What behavior does the female exhibit to discourage the males or show her refusal to
mate?
4. What advantage does the male gain from transferring the sex peptide to the female?
EXERCISE 3: Life Stages and Anatomy
Use the larval cultures provided for you to locate and observe
each life stage in Drosophila development.
1. Transfer any adult flies into an empty fly vial: Gather
an empty vial and stopper from the “Empty Vials”
bin. Place a funnel in the top of the empty vial. Tap
the vial containing flies against the mouse pad, pull
out the stopper, and quickly invert the vial into a
funnel. Tap the vial that now contains flies twice,
remove the funnel, and quickly insert the foam
stopper. Set vial containing adult flies aside.
2. Use the white spatula to remove some blue food
media from the base of the vial and place it on a slide.
Add a drop of Ringer’s solution to the media so the
larvae don’t dry out.
3. Observe your larvae and pupae using the dissecting
scope. Try to identify the different parts using the
following schematic of a third instar larva. (Fat bodies
and circulatory system are not shown for clarity).
QUESTIONS:
1.
What larval organs can you identify in your observations of the 3rd instar? Draw a
diagram of the instars you observe.
NOTE: Not all the organs pictured in the diagram above will be visible in the instars you observe.
2.
Where in the vial is each life stage found? How does this location relate to what its
function is at this point in development?
Hint: What is it doing that it needs to be in that location?
EXERCISE 4: Observing Drosophila Polytene Chromosomes
View the polytene chromosomes on the prepared slides, and answer the following questions:
1. Explain in your own words why the banded pattern on the chromosomes is useful to
geneticists.
2. How would an organism benefit from this many copies of DNA?
3. How does the banding pattern relate to levels of RNA transcription?
EXERCISE 5: Identifying Phenotypes in Wildtype & Mutant Drosophila
Some Drosophila mutations are very obvious while others are subtle and difficult to see.
First, observe wildtype Drosophila adults in the frozen petri dishes provided. Use the
paintbrushes provided to move the flies around, and the forceps and dissecting pins for further
examination. Also observe the live flies. Note particularly:
1. Color of the eye and regularity of ommatidia (an ommatidium is a unit of the compound
eye - each Drosophila eye has 800 ommatidia)
2. Wings and the veins within them, flatness of wings, and overall ovoid shape of wings
3. Bristles on the dorsal thorax, including thickness and length
4. Is a sex comb present or absent on a male foreleg?
Next, identify the mutants provided in the frozen petri dishes. You may take them out of the
dishes for easier observations, but be sure to put them back in the correct dish. Compare your
mutants to the ones pictured in the posters to identify them by name. There are also vials of
live mutants for you to observe.
QUESTIONS:
1.
Can a behavior be a phenotype? Why or why not?
2.
Name each of the six mutants you observed and describe each phenotype: