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Investigating Modes of Inheritance in the Model Organism Drosophila melanogaster

Libby D’Albero
Dr. Tara Luke
Genetics Lab - BIO 2115
30 October 2022
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Abstract
Using the model organism Drosophila melanogaster, I conducted an experimental cross
to determine the mode of inheritance for the eye color gene by observing the progeny across
three generations. The parental generation included the mating of a white-eyed female (XWXW)
and a red-eyed male (XW+Y). Over the course of the next three weeks, the total individuals in the
parental generation (P generation), first filial generation (F1 generation), and second filial
generation (F2 generation) were recorded based on four phenotypic/genotypic categories;
red-eyed males (XW+Y), white-eyed males (XWY), red-eyed females (XW+__), and white-eyed
females (XWXW). By analyzing the category results in the F1 generation using chi-square
analysis, I found my data to support my hypothesis that the mode of inheritance for eye color in
D. melanogaster is X-linked where the allele for white eyes is X-linked recessive and the allele
for red eyes is X-linked dominant.
Introduction
Drosophila melanogaster is a species of fly commonly known as the fruit fly. This
species is a model organism, meaning members are ideal subjects for genetic investigation.
While D. melanogaster has a short generation time, it exhibits a complex life cycle. After just
eight hours from emergence, a female fly is ready to reproduce (Leonelli et al). This species
provides large numbers of progeny from just a singular genetic cross and is easily cultured in a
laboratory environment. Additionally, D. melanogaster has a small genome, just five percent of
the size of the human genome (Pierce, A3). It is well distinguished in appearance and thoroughly
studied by geneticists, and in 2000 the complete species genome was sequenced
(knowyourgenome.org). In this experiment, I conducted a controlled cross between two distinct
genotypes of Drosophila melanogaster and analyzed the progeny of the following F1 and F2
generations. The objective of the experiment was to determine the mode of inheritance of the
present genes in the P generation by comparing the phenotypic ratios found in subsequent
generations using chi-squared analysis. Depending on how closely observed phenotypic values
mimic the expected phenotypic values, a trait may be autosomal dominant, autosomal recessive,
X-linked dominant, or X-linked recessive. Based on the results of the experimental cross, the
mode of inheritance for white eye color in D. melanogaster must be X-linked recessive, whereas
the mode for scarlet, or wild type, eye color must be X-linked dominant.
Methods
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In Week 1, I was assigned one male fly with scarlet eyes (XW+Y) and one female fly with
white eyes (XWXW) from the Wards Biological Supply Co. stock. I placed the flies in an enclosed
vial with a medium of cooked cereal, yeast, mold inhibitor, and color and left the adults to
reproduce. This generation, labeled generation P, reproduced to form the F1 generation.
At the start of Week 2, I tallied the composition of the F1 generation by four categories;
red-eyed males (XW+Y), white-eyed males (XWY), red-eyed females (XW+__), and white-eyed
females (XWXW). Male and female members of D. melanogaster can be visually differentiated
based on sexual dimorphism, where female adults are 10-15% larger than males. Males have
broader black stripes and fewer abdomen segments compared to females and possess visual
distinctions on their front legs called sex combs (course manual).
In order to carry out a survey of the F1 generation, I transferred the flies from the original
vial to a second vial with no medium. After relocating the adult flies, I placed the new vial into
an ice bath in order to anesthetize the subjects for inspection. Once immobilized, I transferred the
flies onto a covered cold pack under a microscope and conducted the tally. All counted flies from
the F1 generation were discarded in ethanol afterwards. The remaining fly stages in the medium
vial were left to produce the F2 generation.
At the start of Week 3, I carried out the same process of anesthetization in order to count
the members of the F2 generation. I tallied the composition by the same four categories;
red-eyed males, white-eyed males, red-eyed females, and white-eyed females. Similarly, all
examined flies were later discarded in ethanol and the remaining fly stages in the original
medium vial were left to develop.
At the start of Week 4, I carried out the same process of anesthetization and tallying of
any remaining members of the F2 generation. All adult flies were discarded in ethanol and any
remaining fly stages in the original medium vial were returned to the instructor for disposal.
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Results

Raw Data

Date of Data Phenotype

Collection Generation Genotype Phenotype Male Female Total

9/13/2022 P W+__ red eyes 1 0 1

9/13/2022 P WW white eyes 0 1 1

Sex Total 1 1

Grand Total 2

9/20/2022 F1 W+__ red eyes 2 10 12

9/20/2022 F1 WW white eyes 15 0 15

Sex Total 17 10

Grand Total 27

9/27/2022 F2 W+__ red eyes 32 27 59

9/27/2022 F2 WW white eyes 31 20 51

10/04/2022 F2 W+__ red eyes 62 70 132

10/04/2022 F2 WW white eyes 73 66 139

Sex Total 198 183

Grand Total 381

Figure 1: Compilation of the fly data collected over four weeks where information is segregated
by generation, sex, genotype and phenotype.
On September 13, the Week 1 grand total for the P generation flies was 2 flies; 1 male
and 1 female. The phenotype of the male was red eyes meanwhile the phenotype of the female
was white eyes. The genes listed under the “Genotype” category were (incorrectly) assumed as
autosomally inherited and are denoted as such. The appropriate genotypes for eye color and
listed correctly in later figures.
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On September 20, the Week 2 grand total for the F1 generation was 27 flies. Of the 27
flies counted, the male:female ratio was 17:10. The red-eye:white-eye ratio was 12:15. More
specifically, there were 2 red-eyed males, 10 red-eyed females, 15 white-eyed males, and 0
white-eyed females.
The totals for the F2 generation were found by summing the results of September 27 and
October 4 (Week 3 and Week 4). The grand total for the F2 generation was 381 flies. Of the 381
flies counted, the male:female ratio was 198:183. The red-eye:white-eye ratio was 191:190.
More specifically, there were 94 red-eyed males, 97 red-eyed females, 104 white-eyed males,
and 86 white-eyed females
F1 Generation (P x P)

White-eyed mother

XW XW

XW+ XW+XW XW+XW

Red-eyed father Y XWY XWY

Figure 2: Diagram of the cross where the two members of the P generation, white-eyed female
and red-eyed male, mate to produce two distinct genotypes.
The theoretical progeny make up of the F1 generation (where two P generation members
are crossed) is described in the above cross diagram. The cross has been made working under the
hypothesis that the mode of inheritance for eye color is X-linked where the allele for white eyes
is X-linked recessive and the allele for red eyes is X-linked dominant. A white-eyed female
(XWXW) crossed with a red-eyed male (XW+Y) will yield a group of offspring where 100% of the
female progeny are red-eyed (XW+XW) and 100% of the male progeny are white-eyed (XWY).
F2 Generation (F1 x F1)

Red-eyed mother

XW+ XW

White-eyed XW XW+XW XWXW

father Y XW+Y XWY

Figure 3: Diagram of the cross where two members of the F1 generation, red-eyed homozygous
female and white-eyed male, mate to produce four distinct genotypes
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Given that all females in the F1 generation were red-eyed and all the males in the F1
generation were white-eyed, a cross diagram of the F2 generation can be produced. A
heterozygous red-eyed female (XW+XW) crossed with a white-eyed male (XWY) will yield a group
of offspring where 50% of the female progeny are red-eyed (XW+XW), 50% of the female progeny
are white-eyed (XWXW), 50% of the male progeny are red-eyed (XW+Y), and 50% of the male
progeny are white-eyed (XWY).

Progeny Results

Expected Percentage (out of Expected Observed

Phenotype Genotype sex total) Count Count

Red-eyed
F1 males XW+Y 0% 0 2

White-eyed
males XWY 100% 17 15

Red-eyed
females XW+__ 100% 10 10

White-eyed
females XWXW 0% 0 0

Red-eyed
F2 males XW+Y 50% 99 94

White-eyed
males XWY 50% 99 104

Red-eyed
females XW+__ 50% 91.5 97

White-eyed
females XWXW 50% 91.5 86
Figure 4: Comparison of the expected progeny genotype percentages and totals versus the
observed progeny genotype percentages and totals segregated by generation.
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I used the resulting ratios in the diagrammed crosses of Figure 2 and Figure 3 to calculate
and compare the expected counts to the observed counts for each phenotype. I calculated the
expected counts by multiplying the percentage for that particular genotype by the corresponding
sex total of that generation (located in Figure 1). Based on the crosses, 0% or 0 males should be
red-eyed in the F1 generation, whereas I actually observed 2 red-eyed males. 100% or 17 males
should be white-eyed, whereas I observed 15. 100% or 10 females should be red-eyed, which
matches the observed 10. Finally, 0% or 0 females should be white-eyes, which matches the
observed 0.
Meanwhile, 50% or 99 males should be red-eyed in the F2 generation, whereas I actually
observed 94 red-eyed males. 50% or 99 males should be white-eyed, whereas I observed 104.
50% or 91.5 females should be red-eyed, whereas I observed 97. Finally, 50% or 91.5 females
should be white-eyed, whereas I observed 86.
Discussion
The data supports my hypothesis where the mode of inheritance for white eye color in D.
melanogaster is X-linked recessive and the mode of inheritance for red eye color is X-linked
dominant. The data for over 400 flies was collected over three generations to determine how eye
color is passed from generation to generation. In the F1 generation, if red eyes are X-linked
dominant and white eyes X-linked recessive, then you would expect the ratio of red male : white
male : red female : white female to be 0:1:1:0. I actually observed a ratio of 2:15:10:0. In the F2
generation, if red eyes are X-linked dominant and white eyes X-linked recessive, then you would
expect the ratio of red male : white male : red female : white female to be 1:1:1:1. I actually
observed a ratio of 94:104:97:86 (Figure 4).
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Observed Expected 2
(𝑂−𝐸)
Phenotype, Genotype (O) (E) 𝐸

Red-eyed male, XW+Y 94 99 0.253

White-eyed male, XWY 104 99 0.253

Red-eyed female, XW+__ 97 91.5 0.331

White-eyed female, XWXW 86 91.5 0.331

2
𝑋 = 1.168
Figure 5: Chi-square analysis of the four categories examined in the F2 generation of
flies.
While the actual ratios and expected ratios seem to correspond upon superficial
inspection, I used the chi-square test to analyze the degree of “agreement” between the observed
and expected values. The hypothesis I looked to confirm was the mode of inheritance for eye
color in D. melanogaster is X-linked where white eyes are recessive and red eyes are dominant.
2
2 (𝑂−𝐸)
The chi-square equation is 𝑋 = Σ 𝐸
where O is the observed value, E is the expected value,

and each term represents one of the four phenotypes (Figure 5).
I chose to conduct chi-square analysis on the F2 generation only. Determining the mode
of inheritance of a trait typically requires observation of at least three generations in order to
associate patterns that may begin to establish. Additionally, using chi-square analysis on my data
for the F1 generation would not be as helpful, considering that the grand total is much smaller
than that of F2.
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Figure 6: Table of values for chi-square analysis (Pierce, 65)

2
For the four F2 phenotypes 𝑋 = 1.168. When there are three degrees of freedom
(df=number of categories - 1) the 0.9 > P > 0.5 (Figure 6). Most scientists assign the cutoff of
0.05, where any P value below 0.05 indicates a statistical significance between the observed and
expected values. The closer P is to the integer 1, the more “fitting” the observed values are to the
expected values. My P value is well above 0.05, meaning the variation in my data is not
statistically significant and any differences can be assigned to chance. So, I may accept the
hypothesis that the mode of inheritance for eye color in D. melanogaster is X-linked where white
eyes are recessive and red eyes are dominant.
Regardless of the acceptance of my hypothesis, possible areas of error exist. A consistent
issue I encountered was accurately trapping and counting my flies. During the transfer from the
medium vial to the empty vial, I lost up to 15 flies every time. Over three weeks of data
collection, it is possible that I lost as many as 40 flies from my generation totals. While more
practice in this technique would reduce this source of error, a more efficient solution would be
finding a safer way to transfer flies.
Another area of interest is my data from Figure 4 where the expected count for red-eyed
males in the F1 generation was 0 and my observed count was 2. Given the cross between the P
generation where a red-eyed male and a white-eyed female mated, it should not be possible to
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produce a red-eyed male. One may attribute the presence of 2 individuals to a mutation, but the
probability of the occurrence of mutation is far too low to produce two individuals out of 27
flies total. Two sources of this phenomenon are possible. First, perhaps a red-eyed heterozygous
female was accidentally introduced to the vial and was able to produce 2 red-eyed males.
However, I more likely incorrectly identified those two individuals as male when they were
female.
I will note that my sex identification abilities progressed as the weeks continued. By
Week 4 I was able to distinguish between the sexes mostly by eye. So this degree of human error
should have a lesser effect as the generations continue.
A continuation of the research conducted in this experiment could entail a replication of
this experiment with larger sample sizes in order to yield more accurate results. An augmentation
of this experiment could involve the breeding of many red-eyed males with many white-eyed
females in the P generation. Some relevant applications of these results could include locating
mutations or identifying particular mutation rates and applying those findings to the human
genome. For example, if the unexpected count of red-eyed males in the F1 generation discussed
previously was actually a result of genetic mutation, we would investigate the mutation further.
Understanding its chromosomal locus and mutation rate could then be matched with potential
human equivalents.
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Works Cited
Leonelli, Sabina, and Rachel A. Ankeny. "What makes a model organism?." Endeavour 37.4
(2013): 209-212.
Pierce, Benjamin A. Genetics: A Conceptual Approach. Palgrave Macmillan, 2012.
“Why Use the Fly in Research?” Knowyourgenome.org, 21 July 2021,
https://www.yourgenome.org/facts/why-use-the-fly-in-research/#:~:text=The%20relation
ship%20between%20fruit%20fly,found%20in%20the%20fruit%20fly.