Lab 1 Lab Manual

This document provides instructions for a genetics laboratory exercise on observing polytene chromosomes from the salivary glands of red worms. Students will dissect and stain salivary glands to view polytene chromosomes under a microscope and learn about their formation and genetic information.

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Lab 1 Lab Manual

Uploaded by

makabigail7

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BIOL2313 Genetics Laboratory 2023-2024

Exercise 1
Observation of polytene chromosome from Chironomus salivary glands

In the insect order Diptera, a unique type of giant chromosome is found which serves as an
excellent material for cytogenetic studies. This chromosome structure is called the polytene
chromosome. Polytene chromosomes were first observed and described by E.G. Balbiani in
1881. They are mostly found in the salivary glands of the dipteran flies: the best understood
are those of Drosophila and Chironomus. The giant chromosomes are the result of endomitosis.
During this process, there is no cell division but only DNA replication. The chromonemata (the
arm of chromosome) replicates without separation. This led to the building up of polyteny (or
multi-strandedness). Furthermore, the two homologues of each chromosome pair remain
permanently synapsed and are considered to be perpetual interphases. Because of this synapsed
nature, chromosome alterations, including inversion, deletions and translocations, can be
readily identified. Besides, a phenomenon called “puffing” is frequently observed in polytene
chromosomes, which is the result of gene activity.

In this lab session, you are going to observe the polytene chromosome from the salivary glands
of the red worm (Chironomus sp.) and experiencing the dissection, staining and preparation of
a slide displaying the polytene chromosome.

Learning outcomes:

After this lab session, you should be able to:

1. Observe the features of a polytene chromosome.
2. Take a photo of a polytene chromosome under microscope.
3. Handle the microscope with care.
4. Dispose different waste according to the safety guidelines.
5. Experience the dissection and staining of salivary glands from red worm with pairs of
forceps.
6. Understand the nature of a polytene chromosome
i). how it is formed and
ii). what information it can provide us with.

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BIOL2313 Genetics Laboratory 2023-2024

Procedures:
1. Place one drop of insect saline solution on a glass slide.
2. Place a red worm from the stock culture in the saline solution on the slide.
3. Dissect the worm with your naked eye or with the stereomicroscope:
i). Use a pair of fine forceps to hold anterior 1/3 of the worm to hold it in place.
ii). Use another pair of forceps to hold the head of the worm GENTLY and SLOWLY pull
the head out of the body.

iii). The salivary glands appear as 2 shortened sausage-shaped bags attaching to the head.

iv). Detach the mouth parts and other associated organs from the salivary glands (if possible).

4. Remove other unwanted tissues from the glass slide.

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BIOL2313 Genetics Laboratory 2023-2024

5. Stain the salivary gland with one drop of aceto-orecin for 10-15 minutes. Place the slide
on the bench on top of a tissue paper.

6. After 10-15 minutes, lower a coverslip over the salivary gland.

7. Put two pieces of blotting paper on top the slip and slightly exert downward pressure to
squash the salivary glands and to soak away the excessed stain.

8. Examine the squashed slide with a compound microscope.

9. Take a photo and label the polytene chromosome you observed.

* Preparation of aceto-orecin stain:

Dissolve 1% orecin in 50% acetic acid. Heat the solution to boiling. Cool and filter. Store in
staining bottles.

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BIOL2313 Genetics Laboratory 2023-2024

Reference figures:

Figure 1. Isolated salivary glands of red worm Chironomus sp. under stereomicroscope.

Figure 2. Salivary glands under staining in a drop of aceto-orecin.

Figure 3. Staining intact salivary gland. Notice the salivary duct, the gland cells and the
nuclei.

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BIOL2313 Genetics Laboratory 2023-2024

Figure 4. Stained salivary gland (left) and the giant polytene chromosome in a gland cell
(right). Notice the nuclei (left) and the polytene chromosome (right).