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Drosophila Crosses

Drosophila Crosses


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I have 2 transgenic lines of Drosophila. I would like to cross them and get both transgene in the same fly. To start this cross, does it matter which fly is female or male? Or as long as the female is virgin, it does not matter?


As long as none of your genes are X-linked and you use virgins it shouldn't matter and you'd get the same proportion of double homozygotes. You may want to perform a reciprocal cross (https://en.wikipedia.org/wiki/Reciprocal_cross) if you don't know what chromosome these genes are on (though flybase.org is useful too).


Drosophila genetics simulation

Developed at the University of Wisconsin-Madison, CGS allows students to perform virtual test crosses with model organisms. Instructors can set the parameters for the populations under study, such as the number and type of traits in a population, the modes of inheritance and trait linkage. Students determine which crosses to perform and interpret the resulting data. CGS can be used as a primary laboratory module for introductory biology or genetics courses, or as a supplement to a hands-on genetics module with real organisms. In addition to mice and Arabidopsis (plants), test crosses can be performed with Drosophila melanogaster (fruit flies) using the CGS software. If you do not have an account and would like to explore some example populations for yourself:

How do I get started?
Click here to launch CGS in a new window.
Choose VIEW PRACTICE POPULATIONS from the main menu.
Choose a population to examine, and decide which test crosses you want to perform.

If you want to explore all of the features of CGS, you can:

Below is a screen capture of a Drosophila population being investigated:

Drosophila flies carry three sets of autosomes and two sex chromosomes. Their relatively fast life cycle and their ease of use made this common fruit fly an important historical model species, and they are still studied by geneticists today. Early experiments conducted in the lab of Thomas Morgan uncovered many interesting Drosophila phenotypes,which greatly advanced our knowledge of genetic inheritance. Students working with CGS are able to perform some of the same experiments that have lead to major breakthroughs in the field of genetics.

For more information about CGS, use the menu on the left to explore the website. If you would like to use CGS for your biology or genetics course, click here to REQUEST AN ACCOUNT.


Resources

Life Science

  • Genetics with Drosophila F1 Crosses

Product Support

  • Carolina Arthropods Manual
  • Living Organism Care Guide: Drosophila

Genetics of Drosophila Melanogaster

Introduction:
Gregor Mendel revolutionized the study of genetics. By studying genetic inheritance in pea plants, Gregor Mendel established two basic laws of that serve as the cornerstones of modern genetics: Mendel’s Law of Segregation and Law of Independent Assortment. Mendel’s Law of Segregation says that each trait has two alleles, and that each gamete contains one and only one of these alleles. These alleles are a source of genetic variability among offspring. Mendel’s Law of Independent Assortment says that the alleles for one trait separate independently of the alleles for another trait. This also helps ensure genetic variability among offspring.
Mendel’s laws have their limitations. For example, if two genes are on the same chromosome, the assortment of their alleles will not be independent. Also, for genes found on the X chromosome, expression of the trait can be linked to the sex of the offspring. Our knowledge of genetics and the tools we use in its study have advanced a great deal since Mendel’s time, but his basic concepts still stand true.
Drosophila melanogaster, the common fruit fly, has been used for genetic experiments since T.H. Morgan started his experiments in1907. Drosophila make good genetic specimens because they are small, produce many offspring, have easily discernable mutations, have only four pairs of chromosomes, and complete their entire life cycle in about 12 days. They also have very simple food requirements. Chromosomes 1 (the X chromosome), 2, and 3 are very large, and the Y chromosome – number 4 – is extremely small. These four chromosomes have thousands of genes, many of which can be found in most eukaryotes, including humans.
Drosophila embryos develop in the egg membrane. The egg hatches and produces a larva that feeds by burrowing through the medium. The larval period consists of three stages, or instars, the end of each stage marked by a molt. Near the end of the larval period, the third instar will crawl up the side of the vial, attach themselves to a dry surface, and form a pupae. After a while the adults emerge.
Differences in body features help distinguish between male and female flies. Females are slightly larger and have a light-colored, pointed abdomen. The abdomen of males will be dark and blunt. The male flies also have dark bristles, sex combs, on the upper portion of the forelegs.

Hypothesis:
After performing a dihybrid cross between males with normal wings and sepia eyes and females with vestigial wings and red eyes, we expect to see only hybrids with normal wings and red eyes in the first filial generation. Then we expect to observe a 9:3:3:1 ratio of phenotypes in the second filial generation.

Materials and Methods:
The materials used for this lab were: culture vial of dihybrid cross, isopropyl alcohol 10%, camel’s hair brush, thermo-anesthetizer, petri dish, 2 Drosophila vials and labels, Drosophila medium, fly morgue.

A vial of wild-type Drosophila was thermally immobilized and the flies were placed in a petri dish. Traits were observed. A vial of prepared Drosophila was immobilized and then observed under a dissecting microscope. Males and females were separated and mutations were observed and recorded. The parental generation was placed in the morgue. The vial was placed in an incubator to allow the F1 generation to mature.
The F1 generation was immobilized and examined under a dissecting microscope. The sex and mutations of each fly were recorded. Five mating pairs of the F1 generation were placed into a fresh culture vial, and the vial was placed in an incubator. The remaining F1 flies were placed in the morgue. The F1 flies were left in the vial for about a week to mate and lay eggs. Then the adults were removed and placed in the morgue. The vial was placed back in the incubator to allow the F2 generation to mature. The F2 generation was immobilized and examined under a dissecting microscope. The sex and mutations of each fly were recorded.

Table 1 Phenotypes of the Parental Generation

Phenotypes Number of Males Number of Females
Normal wings/red eyes 0 0
Normal wings/sepia eyes 3 0
vestigial wings/red eyes 0 4
vestigial wings/sepia eyes 0 0

Table 2 Phenotypes of the F1 Generation

Phenotype Number of Males Number of Females
Normal wings/red eyes 78 95
Normal wings/sepia eyes 0 0
vestigial wings/red eyes 0 0
vestigial wings/sepia eyes 0 0

Table 3 Phenotypes of the F2 Generation

Phenotypes Number of Males Number of Females
Normal wings/red eyes 4 7
Normal wings/sepia eyes 4 5
vestigial wings/red eyes 0 1
vestigial wings/sepia eyes 0 0
normal red/mutated body shape 2 0
normal sepia/mutated body shape 1 0
  1. How are the alleles for genes on different chromosomes distributed to gametes? What genetic principle does this illustrate?
    The alleles on different chromosomes are distributed independently of one another, demonstrating Mendel’s Law of Independent Assortment.
  2. Why was it important to have virgin females for the first cross (yielding the F1 generation), but not the second cross (yielding the F2 generation)?
    It was important to have virgin females for the first cross to ensure that the offspring are the result of the desired cross. It was not necessary to isolate virgin females for the second cross because the only male flies to which they had been exposed were also members of the F1 generation.
  3. What did the chi-square test tell you about the validity of your experiment data? What is the importance of such a test?
    The chi-square test showed that the results of our first cross were valid, but that the results of our F1 cross were not normal. It is important to conduct such a test to determine how much your experimental data deviated from what was expected.

Discussion and Conclusion:
The results of our parental cross turned out just as expected, but our F2 generation was not normal. Some sort of mutation must have occurred that caused the strange body shape seen in several individuals of our F2 generation.


Sex Linked Inheritance: Sex-Linkage in Drosophila and Man (With Diagram)

The chromosomes present in the diploid cells of the majority of the sexually reproducing animals are of two types: autosomes bearing genes for somatic characters and sex chromosomes bearing genes for sex.

Sex chromosomes also carry some genes for non-sexual characters such as colour blindness and haemophilia.

Such genes which are always associated with sex chromosomes are called sex-linked genes. In man and Drosophila the sex chromosomes (X and Y) are unequal in size and shape, X being larger and rod shaped whereas Y is small and slightly curved. In birds and butterflies the sex chromosomes (Z and W) are also unequal in shape and size, Z being larger than W.

In Mendelian pattern of inheritance, the genes for contrasting characters were located on autosomes but not on the sex chromosomes. Secondly, the result of reciprocal cross is same as normal cross which is not the case with sex linked inheritance. There are three types of sex-linked genes depending upon their association with particular chromosome.

(i) The genes which are located on X-chromosomes are called X-linked genes or sex linked genes.

(ii) The genes which are located on Y chromosomes are called Y-linked genes or holandric genes.

(iii) Certain genes are found to occur in both X and Y chromosomes. Such genes are called incomplete sex-linked genes.

In order to understand the inheritance of character present in sex chromosomes, let us understand transmission of X-chromosome from male individual in Drosophila or in man. The X-chromosome from male individual will always pass to the daughter, while X-chromosomes from female individual will be distributed equally among the daughter and sons (Fig. 5.17).

A character from the father goes to the daughter (F1) and then from daughter to grandson in the next generation (F2). Such type of inheritance is also called as criss-cross inheritance. In this type of inheritance result of the reciprocal crosses are not identical as in case with Mendelian crosses.

Sex-Linkage in Drosophila:

T.H. Morgan (1910) for the first time discovered sex-linkage in Drosophila melanogaster. Morgan when experimenting noted the sudden appearance of one white-eyed male (mutant form) in the culture of normal red-eyed Drosophila. This white-eyed male was crossed with red eyed female. The F1 flies (both male and female) were all red-eyed indicating that white eye colour is recessive to the normal red eye colour.

When these F1 flies were inter-crossed freely, the red-and white-eyed flies appeared in the ratio 3: 1 in the F2 generation. White- eyed flies were male. Among the red eyed flies two-third were female and one-third were male. The females were all red eyed whereas 50% males were white eyed and the remaining 50% males were red eyed (Fig. 5.21).

If a reciprocal cross is performed between white eyed female and red eyed male individual, all female individuals in Ft generation are red eyed and all male individuals, are white eyed. When these two types of individuals from F1 generation are inter crossed, female population in F2 generation will consist of 50% red eyed and 50% white eyed individuals. Similarly the male population in this generation consists of 50% red eyed and 50% white eyed individual (Fig. 5.19).

The inheritance of white-eye colour in Drosophila can be explained on the basis of the following assumptions:

(i) Gene for white eye colour in male Drosophila is located in X-chromosome and Y chromosome is empty, carrying no normal allele for eye colour.

(ii) In white eyed female Drosophila there are two X chromosomes, each one bearing a gene for white eye colour (w). It transmits one gene for white eye colour (w) to each offspring.

(iii) As we can see in the above reciprocal crosses, the gene for recessive white eye colour (w) passes by father on to daughter (F1 generation). The daughter in turn passes this gene to her sons (F2 generation). The character thus seems to alter or cross from one sex to the other in its passage from generation to generation. In other words, character is transferred from mother to son and never from father to the son.

Characteristics of Sex Linked Inheritance:

(a) It is a criss-cross inheritance as the father passes its sex-linked character to his daughter who in turn passes it to the grandson.

(b) Daughter does not express the recessive trait but act as carrier in the heterozygous condition.

(c) Female homozygous for recessive trait expresses the trait.

(d) Any recessive gene borne by the X chromosome of male is immediately expressed as Y chromosome has no allele to counteract.

Sex Linkage in Man:

In man about fifty six sex-linked genes have been reported, the most common examples are:

1. Red green colour blindness.

1. Red Green Colour Blindness:

Colour blindness is an example of sex linked character. Those who suffer from red green colour blindness cannot distinguish between red and green colour. The gene for this defect is located on X chromosome. It was first studied by Horner (1876). Colour blindness is recessive to normal vision.

(i) Normal Woman and Colour Blind Man:

When a normal woman is married to a colour blind man, their children (daughters and sons) have normal colour vision. But when their daughters were married to normal man, 50% of their sons are colour blind and the remaining 50% are normal, while the daughters were all normal.

(ii) Colour Blind Woman and Normal Man:

If a colour blind woman marries a normal man, their daughters are normal but all their sons are colour-blind. When these F1 daughters are married to colour blind men, colour blind sons and daughters are born in equal number.

2. Haemophilia (Bleeder’s Disease):

Haemophila is another popular example of sex linked inheritance in human beings. It is caused by a mutant gene (h) present in X chromosome and recessive to normal gene and is, therefore, suppressed in heterozygous condition. Individuals suffering from this disease lack a factor responsible for clotting of blood. So in the absence of blood clotting substance, a minor cut or injury may cause prolonged bleeding leading to death. This disease in man is generally restricted to male members.

If a haemophilic man marries a normal woman, the daughter are all carriers (phenotypically normal but carries haemophilic gene in one on her X chromosome) but sons are normal. Such a carrier daughter, when marries a normal man transmits the haemophilic gene to half of her son (Fig. 5.20). A haemophilic woman is produced only if a carrier woman is married to a haemophilic man.

Haemophilia is also called ‘Royal disease’ as it is found in certain royal families of Europe. Apparently the gene for haemophilia (h) arose as a mutation in a reproductive cell which produced Queen Victoria of England.

Generally Haemophilia is of two types:

It is the most common type and the patient lacks anti-haemophilic factor (AHF).

(ii) Haemophilia B:

It occurs in about 20% of the patients. It is due to lack of plasma thromboplastin component (PTC).


Methods

Transgenic fly model and fly culture

24B-GAL4 was used as the driver of UAS-mediated opsin expression 3 in the heart of Drosophila melanogaster. By crossing UAS-ReaChR or UAS-NpHR-YFP flies with 24B-GAL4 flies, targeted expressions of ReaChR (UAS-ReaChR 24B-GAL4) and NpHR (UAS-NpHR 24B-GAL4) were achieved in cardiac tissue. Wild type (WT) W118 flies were crossed with 24B-GAL4 flies to obtain 24B-GAL4/+ as the control. Details about fly model development and selection are provided in Supplementary Notes and Supplementary Tables 1 and 2.

We prepared fly food by mixing Formula 4–24 (Instant Drosophila Medium Carolina Biological Supply Company), water, and 10 mM all-trans-retinal (ATR) (Toronto Research Chemicals Inc.) solution, which used 200 proof ethanol as the solvent. The ATR concentrations in food used for culturing ReaChR and NpHR flies were 3 mM and 10 mM, respectively, in order to achieve sufficient opsin expressions in the heart. Developed flies were transferred into a vial with the prepared formula and kept in an incubator at 25 °C for

10 h for cross breeding. Flies in the first generation were obtained at the larval, early pupal, late pupal, and adult stages for cardiac control and optical imaging experiments. Normal food was prepared by mixing formula 4–24 and water for culturing WT flies at the same temperature.

Integrated red-light excitation and OCM imaging system

We developed an integrated red LED excitation and OCM imaging system for simultaneous, non-invasive cardiac stimulation and monitoring in Drosophila (Fig. 1a). The OCM system used a portion of a supercontinuum laser source to provide near-infrared light with a central wavelength of 800 nm and a bandwidth of 220 nm 3 . The axial and transverse resolutions of

3.6 μm in tissue were obtained by using the broadband source and a ×10 objective lens respectively to resolve the fly heart structure. A 2048 pixel line scan camera was utilized to detect the interference signal of the back-reflected light from the two arms of the OCM system.

To perform optogenetic control of heart functioning, we integrated a 617 nm LED light source (Migtex Systems, BLS-LCS-0617-03-22) into the OCM sample arm through a dichroic mirror and focused the light to the same point of the imaging beam through the objective lens with a spot size of

2.2 mm, covering the entire heart tube. Control software provided by Migtex Systems allowed tuning of the light intensity. A Labview program was used to control the pulse width (or duty cycle) and frequency of the LED signal, and to synchronize the M-mode image acquisition for the OCM system through a function generator (Agilent 33210 A, Keysight technologies, USA) and a DAQ card (National Instrument USB-6008).

Optical excitation and OCM imaging protocol

The focused excitation light illuminated the entire heart tube in both ReaChR and NpHR flies at each developmental stage. For excitatory optical pacing, power densities of 7.26 mW mm −2 and 3.63 mW mm −2 , and pulse widths of 20 ms and 10 ms were used for larva and early pupa screenings, and late pupa and adult fly screenings, respectively, to identify the flies which could be paced. Flies that could be successfully paced at frequencies 20% higher than their RHRs were selected for experiments. Three pacing pulse trains were applied serially during each OCM measurement, with each pulse train lasting for

4.5 s. Then, 4096 cross-sectional images were acquired in

32 s at the frame rate of

130 frames/s from the posterior segments of the larval and early pupal heart, and the anterior portions of the late pupal and adult heart. M-mode images were acquired from the cross-sectional images, using ImageJ to analyze the time-lapsed heart function with and without red-light stimulations. Control flies were paced and imaged with identical procedures. For all the identified ReaChR flies, we modulated the excitation power densities from 0.46 mW mm −2 to 7.26 mW mm −2 , the pulse widths from 2 ms to 40 ms, and pulse frequencies from 0.5 Hz above the RHRs to higher values until divergence of HR from PR was observed.

Cardiac arrest and inhibitory pacing protocol

To demonstrate restorable cardiac arrest, 10 s of continuous red light was used to identify viable larva, early pupa, late pupa, and adult flies, with an excitation power density of 7.26 mW mm −2 . After discerning viable flies, power intensities were tuned from 0.12 mW mm −2 to 7.26 mW mm −2 to characterize the effect of stimulation power on the cardiac arrest rate for each developmental stage. For inhibitory pacing, duty cycles of excitation pulse trains were modified for individual flies. Pulse frequencies of 2 Hz, 1 Hz, and 0.6 Hz were tested at the excitation power density of 7.26 mW mm −2 . The single heart contractions between adjacent pulses indicated the heart rate at the respective frequencies.

Protocol for study of heart rate recovery

To investigate the heartbeat recovery after cardiac arrest in NpHR flies, we monitored the heart of each early pupa for 60 s through the OCM imaging technique. Before demonstrating cardiac arrest, the minimum excitation power was determined for individual flies by observing the heartbeat during red light excitation. After allowing the heart to beat for 5 s at the RHR, different cardiac arrests were then generated by stimulating the heart for 1 s, 2 s, 5 s, 10 s, or 20 s. To compare the cardiac-arrest-induced cardiac recovery dynamics between different arrest periods, two-sided Student’s t-tests were used for statistical analysis and results were considered statistically significant at p < 0.05.

Statistical analyses and reproducibility

Experiments were repeated independently on multiple flies as mentioned in the text and figure legends. Data were characterized using two-sided Student’s t-tests and represented as mean ± SD.

Reporting summary

Further information on research design is available in the Nature Research Reporting Summary linked to this article.


Introduction

Uniparental transmission of cytoplasmic genetic elements (i.e., mitochondria and chloroplasts) seems to be a general rule in biology. Particularly in animals, mitochondrial DNA (mtDNA) is maternally transmitted. Several mechanisms of prevention of paternal mtDNA transmission have been evolved, such as sperm with no mitochondria, destruction of sperm mitochondria in the newly fertilized egg, and eventual elimination of sperm mitochondria in the embryo by other means (Birky 2001). The variety of the mechanisms that protect uniparental transmission of mtDNA implies that these mechanisms have been evolved independently. It also suggests that there is strong selective pressure for uniparental transmission of mtDNA. The prevailing theory suggests that uniparental transmission prevents the spread of selfish deleterious mutations of cytoplasmic genetic elements in the population (Hastings 1992 Hurst 1992, 1995). This hypothesis is supported by data from several organisms such as yeast (Williamson 2002), Neurospora (Bertrand et al. 1980), and C. elegans (Clark et al. 2012). These studies have shown that mtDNA with large deletions may proliferate faster and outnumber deletion-free mtDNAs. Thus, smaller mtDNA molecules increase their representation in the mtDNA pool within the organism even though they reduce the organism's fitness. However, the validity of this hypothesis has been questioned for Drosophila (Rand 2011).

The advantage of uniparental transmission through the prevention of the spread of deleterious mutations in the population may come at the expense of deleterious mutation accumulation in the mtDNA molecule. This is because uniparental inheritance creates asexual nonrecombining lineages, which accumulate deleterious mutations faster than their sexual counterparts, a mechanism known as Muller's ratchet (Muller 1964 Felsenstein 1974 Gordo and Charlesworth 2000). Several mechanisms have been proposed to explain how mtDNA may overcome Muller's ratchet. These mechanisms include genetic bottleneck, compensatory mutations, back mutations, recruitment of mtDNA copies from the nucleus and recombination [for a review see Loewe (2006)] as well as purifying selection (Stewart et al. 2008). However, apart from recombination, the efficacy of most of these mechanisms on the elimination of Muller's ratchet has not been tested yet either on theoretical or on experimental basis. Recombination remains the main mechanism for elimination of Muller's ratchet.

Infallible uniparental inheritance of mtDNA would in effect eliminate this route of prevention of Muller's ratchet. This is because uniparental inheritance leads to homoplasmy for the maternal mtDNA (heteroplasmy due to point mutations among the mtDNA molecule of the unfertilized egg or due to mutations occurring during the life of the organism is negligible compared with heteroplasmy that could result from biparental inheritance). In a population of identical mtDNA molecules, recombination generates molecules that are identical to themselves and to parental molecules. Thus, prevention of mutation accumulation and eventual mutation accumulation of the mtDNA molecule would be inevitable. Might, then, leakage of paternal mtDNA have evolved because it provided a means for overcoming this limitation?

Despite the wealth of mechanisms promoting uniparental transmission of mtDNA, paternal mtDNA has been observed occasionally in several animal species such as mouse (Gyllensten et al. 1991), Drosophila (Kondo et al. 1992 Sherengul et al. 2006 Nunes et al. 2013), anchovy (Magoulas and Zouros 1993), sheep (Zhao et al. 2004), and human (Schwartz and Vissing 2002). In all these cases, paternal mtDNA is a small minority in the embryo compared with the maternal mtDNA. Leakage of paternal mtDNA in the embryo has been explained as a breakdown of the mechanisms that promote uniparental transmission. Kaneda et al. (1995) observed that paternal mitochondria are eliminated in intra- but not in interspecific crosses in mouse. Sperm mitochondria in mammals are tagged with ubiquitin during spermatogenesis leading to their recognition and subsequent degradation by the proteolytic machinery in eggs after fertilization (Sutovsky et al. 1999, 2000). Based on these observations, Rokas et al. (2003) proposed a conceptual model for paternal mtDNA leakage which allows mtDNA recombination to occur. According to this model, elimination of sperm mtDNA in the fertilized egg involves a reaction between a nuclearly encoded factor that labels the sperm mitochondria and an also nuclearly encoded factor in the egg cytoplasm. The recognition of the label of sperm's mitochondria by the egg factor is nearly perfect in homospecific crosses because they are both encoded by the same nuclear background. In heterospecific crosses, however, the egg factor and the label of sperm mitochondria are encoded from different nuclear backgrounds and the recognition of each other might be less effective allowing the maintenance of some paternal mitochondria in the egg. The recognition of the sperm mitochondria by the egg factor would become less effective, the more distant are the species involved in the cross because more nonshared mutations would have accumulated in the two independently evolving taxa. The model makes two testable predictions. The first prediction is qualitative and suggests that the leakage of paternal mtDNA into the embryo would be more common in interspecific than in intraspecific crosses. The second prediction is quantitative and suggests that the level of leakage would increase as the genetic distance between the hybridizing species increases. In this study, we test these two hypotheses using Drosophila inter- and intraspecific crosses. If the model is correct, we expect that leakage of paternal mtDNA would be higher in inter- than in intraspecific crosses and that we would find more hybrids with leakage in crosses between more divergent Drosophila species. Furthermore, we detected the presence of paternal mtDNA in individual hybrids. The differential presence of paternal mtDNA in male and female hybrids would be good indication that leakage of paternal mtDNA is not a random phenomenon but might be under genetic control.


Drosophila Crosses - Biology

Objective: Students will learn and apply the principles of Mendelian inheritance by experimentation with the fruit fly Drosophila melanogaster. Students will make hypotheses for monohybrid, dihybrid and sex-linked traits and test their hypotheses by selecting fruit flies with different visible mutations, mating them, and analyzing the phenotypic ratios of the offspring.

The image shown below shows a wild-type female fly (left) and a male fly. Recall that "wild-type" refers to the most common or typical form seen in the wild. A + sign is used to denote when a fly displays the wild-type characteristic.

Introduction

Examine the phenotypes available from the left side menu to answer the following questions.

1. Examine the different types of bristles seen in flies. Geneticists use a shorthand labeling system, F = forked. Identify the phenotypes shown:

2. Compare antennae types. How is "aristapedia" different from wild-type?

3. What are different eye colors in fruit flies? Circle the one that is wild-type.

4. Regarding wing size, what is the difference between apterous and vestigial?

5. What are the body colors in fruit flies?

6. Create a mutant fly with any number of variations and mate it with a wild-type fly. How many offspring were wild-type?

7. Mate the offspring of the cross. Use the analyze tab to get more details about the F2 offspring. (The button to "ignore sex" may make counting easier.)

How many wild-type offspring were produced?

How many mutant flies were produced?

Part 2: Monohybrid Crosses

You may realize that choosing a lot of different types of flies make it difficult to analyze inheritance patterns. Your next tasks will focus on analyzing single traits within flies to determine how they are inherited.

1. Reset all flies in the design tab.
2. Design a male fly with vestigial wings and cross it with a wild-type female
3. Add the results to your "Lab Notes."
4. Mate the offspring of this cross.

5. Based on these two crosses you probably have an idea about how vestigial wings are inherited.

Is VG recessive or dominant?

How do you know?

6. In genetics, numbers are statistically analyzed. The fly simulator has a built into it. Under the Analyze tab, you can click on "Include a test hypothesis."

If your hypothesis that VG is a recessive trait is correct, then you would expect what proportion of the F2 offspring to have vestigial wings?

What proportion would have wild-type wings?

7. Place the expected numbers in the hypothesis field and click on "test your hypothesis." The program will do the chi square calculations.

What is your chi-squared test statistic?

Compare this to the chi square table to determine a goodness of fit.

8. Summary: Explain how vestigial wings are inherited in fruit flies (claim) and provide evidence from your data and chi-square statistic analysis.

Part 3: Sex Linked Traits

1. Cross a white eyed male with a wild-type female.

How many of the offspring are males / red eyes?

How many females / red eyes?

2. Predict what would happen if you crossed two of the offspring. Explain your reasoning by showing a punnett square

3. Perform the cross and use the statistical analysis tool to test your prediction.

4. Summary: Explain how red/white eye color is inherited in fruit flies (claim) and provide evidence from your data and chi-square statistic

Part 4: Lethal Alleles

Aristapedia is a lethal allele that is also dominant. Individuals with this trait must be heterozygous (Aa) because the homozygous condition (AA) is lethal. This is not a sex-linked trait. Wild-type flies do not carry the allele for aristopedia (aa).

1. Predict what the outcome of a cross between a wild-type fly and one with aristopedia. Show the punnett square to illustrate your reasoning.

2. Perform the cross and determine if your prediction is correct using statistical analysis. Summarize your results and indicate whether your prediction is confirmed.

Part 5: Linkage Groups

When two alleles are located on the same chromosome they are inherited together. However, crossing-over can occur during meiosis and the alleles are switched. Vestigial wings (VG) and Black body color (BL) are located on chromosome 2.

1. Cross a female VG, BL fly with a wild-type male. (ggbb x GGBB)

How many wild-type offspring are produced?

What is the genotype of these offspring?

2. Choose a female from the offspring and mate it with a male that has vestigial wings and a black body. Show a punnett square or a visual representation of the alleles involved in this cross to make a prediction about the offspring.

3. Complete the table (ignore sex).

Phenotype Observed Proportion
+ (wild-type)
Vestigial wings (gg)
Black body (bb)
VG, BL (ggbb)

4. How does crossing-over affect the observed outcomes? Explain why the observed flies do not match your prediction.

5. The percentage of crossing over events is used to develop a map of chromosomes. View the chromosome map.

How far apart are the alleles for black body and vestigial wings?

View the proportion of flies from your data that indicate crossover occurred (VG and BL flies) and multiple it by 100. Based on your data, how far apart are these alleles?

Resources

/>This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.


Sex-Linked Traits

Eye color in Drosophila, the common fruit fly, was the first X-linked trait to be identified. Thomas Hunt Morgan mapped this trait to the X chromosome in 1910. Like humans, Drosophila males have an XY chromosome pair, and females are XX. In flies the wild-type eye color is red (XW) and is dominant to white eye color (Xw) (Figure 15). Because of the location of the eye-color gene, reciprocal crosses do not produce the same offspring ratios. Males are said to be hemizygous, in that they have only one allele for any X-linked characteristic. Hemizygosity makes descriptions of dominance and recessiveness irrelevant for XY males. Drosophila males lack the white gene on the Y chromosome that is, their genotype can only be XWY or XwY. In contrast, females have two allele copies of this gene and can be XWXW, XWXw, or XwXw.

Figure 15: In Drosophila, the gene for eye color is located on the X chromosome. Red eye color is wild-type and is dominant to white eye color.

In an X-linked cross, the genotypes of F1 and F2 offspring depend on whether the recessive trait was expressed by the male or the female in the P generation. With respect to Drosophila eye color, when the P male expresses the white-eye phenotype and the female is homozygously red-eyed, all members of the F1 generation exhibit red eyes (Figure 16). The F1 females are heterozygous (XWXw), and the males are all XWY, having received their X chromosome from the homozygous dominant P female and their Y chromosome from the P male. A subsequent cross between the XWXw female and the XWY male would produce only red-eyed females (with XWXW or XWXw genotypes) and both red- and white-eyed males (with XWY or XwY genotypes). Now, consider a cross between a homozygous white-eyed female and a male with red eyes. The F1 generation would exhibit only heterozygous red-eyed females (XWXw) and only white-eyed males (XwY). Half of the F2 females would be red-eyed (XWXw) and half would be white-eyed (XwXw). Similarly, half of the F2 males would be red-eyed (XWY) and half would be whiteeyed (XwY).

Figure 16: Crosses involving sex-linked traits often give rise to different phenotypes for the different sexes of offspring, as in the case for this cross involving red and white eye color in Drosophila. In the diagram, w is the white-eye mutant allele and W is the wild-type, red-eye allele.

Discoveries in fruit fly genetics can be applied to human genetics. When a female parent is homozygous for a recessive X-linked trait, she will pass the trait on to 100 percent of her male offspring, because the males will receive the Y chromosome from the male parent. In humans, the alleles for certain conditions (some color-blindness, hemophilia, and muscular dystrophy) are X-linked. Females who are heterozygous for these diseases are said to be carriers and may not exhibit any phenotypic effects. These females will pass the disease to half of their sons and will pass carrier status to half of their daughters therefore, X-linked traits appear more frequently in males than females.

In some groups of organisms with sex chromosomes, the sex with the non-homologous sex chromosomes is the female rather than the male. This is the case for all birds. In this case, sex-linked traits will be more likely to appear in the female, in whom they are hemizygous.


Biology

The fruit fly, Drosophila melangaster, was used as an instrument to study the inheritance and transmission of some characters. The characters used were eye color (red or white) and antenna mutation. Monohybrid crossings revealed phenotypic ratios of 3 1, supporting the experiments hypothesis.

Introduction
In the early twentieth century, the tiny fruitfly, Drosophila melanogaster, became the wonder geneticists tool. It was made famous by Thomas Morgan, an American geneticist in 1912, who used it extensively in his researches to verify the assertions of Gregor Mendel, and also to locate the position of genes on chromosomes (chromosome mapping).

Drosophila melanogaster, the fruit fly is a very suitable instrument for genetic studies. One of the advantages is that it is tiny, the adult is only 0.5cm long, and so can be kept in the laboratory in large numbers. Drosophila are small flies, typically pale yellow to reddish brown to black, with red eyes (Drosophila, 2009). The males can easily be distinguished from the females because the males have rounded abdomens while the females have pointed abdomens. Also, it completes its life cycle within two weeks and breeds in large numbers, enabling geneticists to follow the transmission of characters through several generations in a short period. It has only homologous pairs of fairly large chromosomes in its somatic cells and it has many easily distinguishable discontinuous characteristics.

Drosophila has been found to have four pairs of chromosomes a pair of sex chromosomes (X or Y) and three pairs of autosomes (2, 3 and 4). The size of the genome is about 165 million bases and contains an estimated 14,000 genes (by comparison, the human genome has 3,300 million bases and may have about 70,000 genes yeast has about 5800 genes in 13.5 million bases) (Introduction to Drosophila melanogaster, 2006). The analysis of the entire genome of the fruit fly has almost been completed.
The aim of this experiment is to verify the hypothesis which states that if there were 10 wild males and 10 mutated females in the F1 generation, there would be a phenotypical ratio of 3 mutated flies to 1 normal fly.

Methods
The materials used during this experiment were adsorbent wand, petri dish, several Drosophila vials and labels, FlyNap solution, fly morgue.
The whole experiment was conducted over the space of three weeks. The procedures will be divided according to the weeks.
Week 1
The first part of the experiment was to anesthetize the flies. This was done using a FlyNap. A wand was prepared and dipped inside the FlyNap solution. The wand was then introduced into the container containing the selected flies and held there for about two minutes. The anesthetized flies were shaken out on a white card. With a dissecting microscope, the adults were observed and their genders and physical characteristics were noted. The adults were separated according to their genders with the use of the properties differentiating the males from the females.
Week 2
The adults were allowed to mate the previous week and the offsprings observed the following week. The fly vials were retrieved and the eggs and larvae checked for. All the flies present were anesthetized using the FlyNap solution. The P generations of flies were removed and the eggs and larvae left in the vial. These were then allowed to incubate in the incubator, pending their observation the following week.
Week 3
A fresh vial of food was prepared for the new generation of flies. Yeast was also added to the food. The flies were labeled with the corresponding mutant letter. Again, the flies were anesthetized using FlyNap. Their characteristics were observed and recorded. The flies were sorted according to their phenotypic characteristics. Of these, 10 males and 10 females were selected and put in the new fresh vials and allowed to mate and produce the F2 generation of offsprings. After some days, the F2 offsprings were observed and their phenotypes recorded (Carolina Drosophila Manual, 2009).

Results
SHAPE MERGEFORMAT
Figure 1. A monohybrid cross between a wild type male and a mutated female.
SHAPE MERGEFORMAT
Figure 2. A second monohybrid cross between a mutated male (antenna) and a White eyed female.
Mutation A represents antenna formation, mutation B represents tan body and white eye, while mutation C represents wingless flies.
After the WB X A monohybrid cross, it was discovered that all the F1 generation of offsprings looked alike, that is all the phenotypes were of the wild type.
Observed of flies 21 males and 12 females.
During a second cross between AB and B flies, all the offsprings expressed the same mutated antennae trait. 20 offsprings had red eyes and 11 others had white eyes.
Observed of flies 8 males and 23 females.

BXbAXbaYAYaXBA Red eyes
XBXbAA
Mutated antenna Red eyes
XBXbAa
Mutated antennaB Red eyes
XBYAA
Mutated antennaB Red eyes
XBYAa
Mutated antennaXBa Red eyes
XBXbAa
Mutated antenna Red eyes
XBXbaa
Normal antennaB Red eyes
XBYAa
Mutated antennaB Red eyes
XBYaa
Normal antennaXbA White eyes
XbXbAA
Mutated antenna White eyes
XbXbAa
Mutated antennaB White eyes
XbYAA
Mutated antennaB White eyes
XbYAa
Mutated antennaXba White eyes
XbXbAa
Mutated antenna White eyes
XbXbaa
Normal antennaB White eyes
XbYAa
Mutated antennaB White eyes
XbYaa
Normal antennaFigure 3. Punnett square with a dihybrid cross showing inheritance patterns through the first and second filial generations of two characters.

Discussion
The monohybrid cross illustrated in Fig. 2 supports the hypothesis. The females which expressed the same phenotype as the parents were all mutant, that is, they had antenna mutation. During the course of the experiment, the cross between WB and A did not work. This was probably due to excess light. All the offsprings had the wild type phenotype. The correct phenotypic ratio should have been ratio 31 of red to white. This would have confirmed that the red eye trait is dominant to the white eye trait. Analysis of why this did not work is beyond the scope of this experiment.

A second cross between AB and B gave a different result. The observed phenotypes were 15 red eyed females with mutated antenna, 8 white eyed females with mutated antenna, 5 red eyed males with mutated antenna and 3 white eyed males with mutated antenna. This would give a phenotypic ratio of 20 red eyed offsprings to 11 white eyed offsprings, although, all had mutated antennae. The observed F1 phenotypes did not match with the expected F1 phenotypes. This is probably due to some errors during the experiment.
Figure 3 shows the results of the dihybrid cross to study the inheritance patterns of the two characters, that is, eye color and antenna mutation. Each member of the F1 generation undergoes meiosis to produce six kinds of gametes, XBA, XBa, XbA, Xba, YA and Ya. It can be seen that the alleles for the two characters were independently assorted. If members of the F1 generation were then allowed to cross among themselves, the F2 generation so produced shows 4 phenotypes and 12 genotypes. The four phenotypes which appear in the ratio 6 2 6 2 are as follows
six red eyed with mutated antenna
two red eyed with normal antenna
six white eyed with mutated antenna
two white eyed with normal antenna
Observing the Punnett square closely, it is noticed that the phenotypic ratio of red eyes to white eyes for each sex was 1 1, that is, the male flies had a 1 1 ratio for red eyes versus white eyes and the female flies, 1 1 ratio for red eyes versus white eyes. Again, the ratio of mutated antenna to normal antenna was 3 1 for all the red eyed males. Same thing can be noticed for all the red eyed females, white eyed males, and white eyed females.


Watch the video: Genetic cross to perform knockdown using UAS-GAL4 system (May 2022).