Genetic Map of Drosophila via Three Point Cross

Introduction

Drosophila mature through complete metamorphosis, as do all members of the order Diptera. Diptera are commonly known as (true: having two wings) flies and include many familiar insects such as mosquitoes, black flies, midges, fruit flies, and house flies. Drosophila also have a small number of chromosomes: three autosomal pairs and X/Y chromosomes. This helps to simplify genetic mapping and study.

Materials and Methods

To complete the experiment a population of virgin males and virgin females of each mutation to be tested was obtained. Each mutation being tested would have its own culture tube with a food source at the bottom. The food source can be any number of things, but for this experiment an instant Drosophila medium was mixed water and a small amount of dry yeast was added. After reproducing, the yeast would serve as a food source. The food source should take up no more than one inch at the bottom of the culture tube. A separate culture tube should be used as a sleep chamber to anaesthetize Drosophila before characterizing the phenotypes.

For each mutation investigated, a culture tube was prepared by the instructor and the flies in these containers served as the F1 generation, the result of parental crossing of wild type males and mutant females. The first step was to anaesthetize the Drosophila so they could be counted and sorted by phenotype. Drosophila can easily be anaesthetized and manipulated with unsophisticated equipment. In the case of this particular experiment the substance provided to anaesthetize the Drosophila with was called FlyNap. FlyNap consists of a mixture of: 50% Triethylamine, 25% Ethanol, and 25% Fragrances. The flies were transferred from the food chamber to the sleep chamber. Once in the sleep chamber, a foam stopper was placed in the tube and a swab dipped in FlyNap was inserted into the container. After roughly 60-90 seconds, the flies were fully anaesthetized. Once anaesthetized, the flies were carefully poured onto a white index card and divided into four categories: male (?) wild type, male (?) mutant, female (?) wild type, and female (?) mutant. A soft, fine-tipped paint brush was used to separate the flies into the appropriate categories to avoid any damage to the specimen. As stated previously, males can be distinguished from females because of a few different characteristics: females are slightly larger than males, having seven bands on the posterior end of their abdomens and males have a dark pigment concentration on their posterior end; males can also be identified by sex-combs on their forelegs. After separating, five males and ten females were placed into new culture tubes, containing a food source. The selection of males and females, as instructed, included both wild type and mutants (if any were present). This was repeated for each mutation and the tubes were labeled and stored for two weeks.

At the end of the two week period, it was discovered that the F2 generation had not emerged from the pupal casing, probably due to a ...