Getting To Know The Flies Better

On Day 4 we had to dissect the fruit fly larvaes in order to look at their polytene chromosomes. It was a totally new and cool experience and we realised that it was not as easy as it seemed. We actually had to "pull" the larvae apart using a disecting needle and a focep to get its salivary glands which contain the polytene chromosomes. It was really difficult to do that as we encountered problems like locating the larvae's head as well as the pinning down of the head and the removal of its body. We failed many times as we were either not able to locate the salivary glands or we were unable to separate the salivary glands from the other body parts. Even though we failed to extract the chromosomes in the end, we learnt about the importance of perseverance and not giving up no matter how tough the challenge is.

Procedures for Polytene chromosomes extraction and staining

1. Remove a larvae from the vial provided and place it onto a drop of 0.7% saline solution
   
2. Using a sharp forcep, a sharp needle, dissect the larvae and look for the salivary glands as shown in the pictures below.


3. Tease out all other tissues and take only the salivary glands.


4. Drain the saline off the slides and replace with HCL( Fixation of chromosomes; preserve the integrity of the structure) for a minute.


5. Drain the HCL and replace with aceto-orcein stain(stain the cells). Let it stand for 10 minutes.
   
6. Drain the stain and replace with acetic acid(fill the stain permanently). Let it stand for 1-3 minutes.


7. Drain and replace with saline.


8. Place a coverslip over the glands, using the thumb and a paper towel, push down on the slide. The pressure applied will squash the glands, rupture the nuclear membrane and free the chromosomes.
   
9. Using a compound microscope, observe the slide at the different magnifications to check for the stained chromosomes.


10. Make the slide permanent by brushing along the edges of the coverslip with nail polish.

More on Polytene Chromosomes

How are they formed?

Polytene chromosomes are formed when some specialized cells undergo repeated rounds of DNA replication without cell division (endomitosis), producing chromatids that remain synapsed together in a haploid number of chromosomes.

Characteristics

They have light and dark banding patterns which can be used to identify chromosomal rearragements and deletions.

Functions

They can increase the volume of the cells nuclei, cause cell expansion and polytene cells may also have a metabolic advantage as multiple copies of genes permits a high level of gene expression.

Taken from http://en.wikipedia.org/wiki/Polytene_chromosome

Banded chromosomes from http://biology.clc.uc.edu/fankhauser/Labs/Genetics/Drosophila_chromosomes/Drosophila_Chromosomes.htm