Drosophila Genetics Homework Packet

Gregor Mendel created two main laws prior to his experiments with his growing pea plants. The first law he created is the law of segregation. It states that the two members of a gene pair (alleles) segregate (separate) from each other in the formation of gametes. Half the gametes carry one allele, and the other half carry the other allele.[1] And the second, which is the law of independent assortment. It states that genes for different traits assort independently of one another in the formation of gametes. These laws were discovered and formed using only pea plants. He used peas because peas are normally self-pollinating, but in this case Mendel could control whether they self-pollinated or not. In his initial experiments, Mendel removed the stamens from some of the young flowers so that self-fertilization could not occur, then tied a bag around each flower so that cross-fertilization could not occur. After the pistils became mature, he artificially cross-pollinated them by dusting the stigma of pea plants with pollen of other pea plants that had factors for some contrasting trait.

In this experiment, Drosophila melanogaster, fruit flies, will be used instead of pea plants. D. melanogaster continues to be widely used for biological research in studies of genetics, physiology, microbial pathogenesis and life history evolution. It is typically used because it is an animal species that is easy to care for, has four pairs of chromosomes, breed quickly, and lays many eggs. D. melanogaster is a common pest in homes, restaurants, and other occupied places where food is served.[2]

Hypothesis

If we believe that the law of dominance and segregation is accurate, then if we mate a red eyed female with a white eyed male, the characteristics would properly cross over to the offspring.

Materials

  • Fruit
  • Two transparent plastic tubes
  • Food substance that can be put in tubes
  • Magnifying lens

Procedure

  • Leave a fruit out in the open for 5 days. Two types of fruit flies should start to appear day by day: flies with white and red eyes. These are the only traits tested.
  • Isolate one red eyed female virgin fly and one white eyed male fly in a suitable mating capacity which can be a tube with appropriate conditions.
  • When the larvae have been created, take out the parents to prevent mating with the first generation. This would convolute the experiment if the parents were unnoticeably allowed to mate with the F2 generation.
  • When the flies have grown up freeze them to immobilize them for a little while. This doesn’t harm them, it just freezes their wings for a little while.
  • Then take them out onto a clean surface by gently shaking the tube downward. Make sure they are all out because each and every one of the flies can alter the outcome of the results.
  • Separate the F2 generation by eye color and gender. Make a chart or a list that shows the exact number of flies in each gender group and separate them again with the color of their eyes.
  • Analyze the results. (Why did this come out the way it did? What were the key factors that influenced the outcomes?)
  • Then mate a male with a female from the F2 generation in a closed tube with food substances inside. Make sure that they are only from the F2 generation.
  • After the female lays her eggs remove the parents from the tube to thwart future mating with the F3 generation.
  • Wait about 14 to 21 days for the flies to grow into full adults.
  • Once the larvae have grown into adults, freeze the tube again to immobilize them for a while.
  • Then separate them again by gender and eye color as before.

Observations

  • Male flies have a small black comb like fragment on their legs unlike the female fly.
  • Female flies a visibly bigger than male flies.
  • The flies in the F2 generation were all red eyed but were not all female
  • After the dihybrid cross, there started to be more white flies.

Analysis

It was observed a small but discrete variation known as white-eye in a single male fly in one of the bottles. I bred the fly with normal (red-eyed) females. All of the offspring (F1) were red-eyed. Brother–sister matings among the F1 generation produced a second generation (F2) with some white-eyed flies, all of which were males. To explain this curious phenomenon, I developed the hypothesis of sex-limited—today called sex-linked—characters, which I assumed were part of the now called X-chromosome of females. Other genetic variations arose in my stock, many of which were also found to be sex-linked. Because all the sex-linked characters were usually inherited together, I gradually became convinced that the X-chromosome carried a number of discrete hereditary units, or factors. I adopted the term gene, which was introduced by the Danish botanist Wilhelm Johannsen in 1909, and concluded that genes were possibly arranged in a linear fashion on chromosomes. Much to my credit, I rejected my skepticism about both the Mendelian and chromosome theories when I saw from two independent lines of evidence—breeding experiments and cytology—that one could be treated in terms of the other.[3]

White Gene, abbreviated w, was the first sex-linked mutation ever discovered in the fruit fly Drosophila melanogaster. I collected a single male white-eyed mutant from a population of Drosophila melanogaster fruit flies, which usually have dark brick red eyes. Upon crossing this male with wild-type female flies, I found that the offspring did not conform to the expectations of Mendelian inheritance. The first generation (the F1) produced 1,237 red-eyed offspring and three white-eyed flies, all males. The second generation (the F2) produced 2,459 red-eyed females, 1,011 red-eyed males, and 782 white-eyed males. Further experimental crosses led me to the conclusion that this mutation was somehow physically connected to the “factor” that determined gender in Drosophila. I named this trait white gene, now abbreviated w.

This is where the now called Y chromosome was introduced. The Y chromosome is one of two sex chromosomes in mammals, including humans, and many other animals. The other is the X chromosome. Y is the sex-determining chromosome in many species, since it is the presence or absence of Y that determines male or female sex. In mammals, the Y chromosome contains the gene SRY, which triggers testis development. The DNA in the human Y chromosome is composed of about 59 million base pairs. The Y chromosome is passed only from father to son, so analysis of Y chromosome DNA may thus be used in genealogical research.

Conclusion

First, I crossed the white-eyed male he had found to a normal female, and I looked to see which trait was dominant in the F1 generation: all the progeny had red eyes. Now, would the white-eye trait reappear, segregating in the F2 progeny as Mendel had predicted? In the F2, there were 3470 red-eyed flies and 782 white-eyed flies, roughly a 3:1 ratio. Allowing for some deficiency in recessives, this was not unlike what Mendel’s theory predicted. But in this first experiment, there was a result that was not predicted by Mendel’s theory: all the white-eyed flies were male! At this point, I had never seen a white-eyed fly that was female. The simplest hypothesis was that such flies were inviable (this might also explain the deficiency of recessives in the 3:1 ratio above).

If any of the F2 females carried the white-eye trait but did not show it, then it should be revealed by a test cross to the recessive parent. It was. Crossing red-eyed F2 females back to the original white-eyed male, I obtained 129 red-eyed females, 132 red-eyed males, and 88 white-eyed females, 86 white-eyed males. Again, this was a rather poor fit to the expected 1:1:1:1 ratio due to a deficiency in recessives. The important thing, however, was that there were fully 88 white-eyed female flies. Clearly, it was not impossible to be female and white-eyed. Why, then, were there no white-eyed females in the original cross?[4]

Bibliography

Garland, E. A. (2014, March 18). Thomas Hunt Morgan, 2.0. Retrieved January 3, 2015, from Encyclopeadia Britannica: http://www.britannica.com/EBchecked/topic/392256/Thomas-Hunt-Morgan/5014/The-work-on-Drosophila

Pearson Education. (2008, June 7). Concept 1: Reviewing Mendel’s Laws. Retrieved from BioCaoch Activity: http://www.phschool.com/science/biology_place/biocoach/inheritance/laws.html

Peterson. (2006).

Wikepedia: The free Encyclopedia. (2014, December 31). Drosophila melanogaster. Retrieved January 2, 2015, from Wikepedia: http://en.wikipedia.org/wiki/Drosophila_melanogaster

[1] (Pearson Education, 2008)

[2] (Drosophila melanogaster, 2014)

[3] (Garland, 2014)

[4] (Peterson, 2006)

 

 

Project

Throughout this section, reference will be made to worksheets given to the students to guide them through the research process with their Drosophila melanogaster crosses.  These sheets may be accessed by clicking on the link provided here or by scrolling down to "Support Materials" below, where they are stored in the order that we used them.

Introducing Drosophila melanogaster 

Several weeks prior to the start of the unit, I (Molly) ordered the flies (wild type, vestigial wing mutant, and white-eye mutant) and their supplies from Carolina Biological supply.  Shortly after they arrived,  I gave the students their first introduction to the flies and asked them to make expansion vials so that we could increase the number of flies as instructed by the Carolina Drosophila manual.

View InstructionsforDrosophilaMedium022706.doc

In addition to having students research basic information such as classification and life cycle from their textbook and the Drosophila manual, I also asked them to perform an initial assessment of their groups' strengths and weaknesses as well as their availability to come in before or after school to count flies when needed.

View IntrotoDrosophiliamelanogaster022706.doc

View DrosophilaGroupStrengthsWeaknesses022706.doc

The final piece of introduction to Drosophila was a worksheet the students completed on fly phenotypes that familiarized them with genetic notation as well as the appearance of the wild-type and mutant flies.

View Phenotypes030206.doc

Students were also asked to draw the various fly phenotypes.

 

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Vestigial wing vs. Wild Type © 2006 fivenavy1

 

 

Click on an image to view larger version & data in a new window


Red-Eyed (Wild Type) vs. White-Eyed © 2006 threeyellow3

 

Male or Female Flies?

In order to make sure students understood the genetic basis of sex as related to sex chromosomes, I asked them to answer some questions based on reading from their biology textbook as well as to take lecture notes as I spoke briefly about this topic.

View SexChromosomes030306.doc

I also wanted to make sure they had the more practical skill of telling male and female fruit flies apart, as this is critical for crossing the flies successfully. Students completed the worksheet below and also did a "practice run" for fly sexing, learning how to use the Fly Nap and then sorting the sleeping flies into male and female piles. 

View MaleandFemaleFruitFlies030606.doc

Students were also asked to draw the difference between male and female flies. This was usually a drawing of fly abdomens copied from the Carolina Drosophila manual, but sometimes students got more creative.

 

Click on an image to view larger version & data in a new window


© 2006 fruitflygreen4

 

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Female fly on the left, Male on the right © 2006 twored3

 

Introducing the Tree of Life

After students had been exposed to basic information about the flies, I, with Kathryn's help, introduced the Tree of Life website to students.  Kathryn did a short presentation on the site and provided students with a paper to keep track of their username and password when they logged into the site.  I had made colored folders for the students in each group to help them keep their fly materials together, so we used their class period and folder color to assign usernames in the ToL (for example, the 2nd period "Red" group would be twored and its members would be twored1, twored2, twored3, and twored4).  

View IntroductiontotheTreeofLifeNotesandInstruct031006.doc

Making the Crosses - Parent and F1 Generations

After an appropriate number of wild type, vestigial and white-eyed flies had grown to maturity, students performed their parent crosses. 

 Wild Type Flies
Vestigial Wing Flies
White-Eyed Flies
 

Click on an image to view larger version & data in a new window


© 2006

 

 

Click on an image to view larger version & data in a new window


© 2006

 


Click on an image to view larger version & data in a new window


© 2006

 

 

 

Before actually putting flies in vials, they completed this sheet so they would have a record of what they did.

View ParentCross031306.doc

Also in this time frame, I asked students to complete an initial self-check of their group folders to make sure each person had the appropriate worksheets completed.

View FlyFolderIndividualWorkGradingCheck1.doc

Approximately a week later, after the parent generation flies had grown to maturity, students made their F1 predictions (drawing punnett squares to illustrate their predictions if the trait of interest to them (vestigial wings or white eyes) was autosomal or sex-linked.

View F1Predictions032106.doc

Providing More Guidance for Website Content 

Kathryn developed a checklist for items to include in the Treehouse website.  We ended up not using a few of the items (for example, an audio recording), adding a few things (the F1 Outcomes and Eulogies the students wrote for their flies)  and changing others (a chi-square analysis became a percent-error calculation) but the checklist helped students stay on task when they were making their websites.

View ChecklistContentforEachGroupsWebpage032006.doc

 Kathryn also conducted a short lesson with students, asking them to think about what makes a good website and giving them some instruction on how to format their sites.

View WEBSITEDESIGN032706.doc

Midpoint Group Reflection 

Around this time, I also asked the student groups to assess themselves on how they were doing as a group.  Students completed a 2-page assessment that asked them what was working in their groups, what was not working, and how they thought they could improve.

View MidpointGroupReflection032006.doc

Outcomes from the F1 Generation and Predicting F2

Students had spent a few days setting up their websites, but without the rest of the experiment, they wouldn't say much.  Approximately one month after we started the fly experiments, students napped their F1 generation and observed their phenotypes.

 

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Flies in a Nap vial © 2006

View Flies Ready to Be Napped

 (NOTE: This is a 13-second clip that takes about 2 mins to download on DSL)

 

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Sorting flies by sex and observing phenotypes© 2006

Students used this sheet as they made their observations to  note the phenotypes and probably genotypes of their flies before  crossing F1 males and females in two new vials to create the F2 generation.

View F1Outcomes033106.doc

A few days later, students made predictions about their F2 outcomes.

View F2Predictions040306.doc

Self Checking Fly Folder Work

Several days after students made their F2 predictions, I asked them to go through their group folders and make sure each student had the proper worksheets completed.

View FlyFolderIndividualWorkGradingCheck2.doc

F2 Outcomes and Percent Error Calculations

Close to two months after starting the fly genetics unit, students had almost completed their experiments. Students napped and counted their F2 generation flies every other day for approximately 1.5 weeks (generally 3 to 4 distinct counting occasions).  They kept track of how many flies of each phenotype they counted using this form.

View F2Outcomes041706.doc

Once they completed their counting, students used this form to calculate their percent error for each of their two vials and assess whether their initial hypothesis about their mutation (it was autosomal, it was sex-linked) was correct.  They also completed several questions that led directly into the conclusions they drew on their group webpages.

View ComparisonsPercentError042506.doc 

Completing the Tree of Life Website

Students spent 3 to 4 (50-minute) class periods finishing their websites.  I helped them with ideas for what to include in the introduction and conclusion by providing them with these instructions:

View SuggestionsforWebsiteIntroandConclusion042706.doc

Final Notes

As some of my students were completing my biology course for Honors credit, I set up a set up a series of questions that students answered about the "Honors Fly Crosses" where flies with multiple mutations were crossed (for example, vestigial wing, white-eyed) and students had to make more complex predictions and do more fine-grained sorting of phenotypes. 

View HonorsFruitFlyQuestions.doc

Near the end of the project, Kathryn set up a PowerPoint presentation to show parents and community members what students had been doing as part of the fruit fly genetics unit. 

View FruitFlyGeneticsPictures.ppt

 

Evaluation

I created a rubric that took into account whether students included all required sections in their page and paid particular attention to their introduction and conclusion to demonstrate their understanding of the project and what they had learned by the end of the experiment. I also awarded points to students who took time to format their web submission in a professional way. Please see the rubric under "Support Materials" below.

Portfolio Pages

Fruit Fly Genetics at City High School
© 2006 fruitflyyellow2
View the outcome of students' projects in their treehouse webpages! This portfolio treehouse contains links to students' investigations.

Information on the Internet

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