Drosophila melanogaster Fruit fly Report

Abstract
This lab involved a monohybrid reciprocal cross and a sex-linked cross utilizing Drosophila melanogaster fruit flies. After sexing the flies, placing them in media, and letting them cross breed, we analyzed and recorded the phenotypes that were observed. The P1 flies were anesthetized then sexed by observing their phenotypes and equally distributed into the media we made for their copulation. When a sufficient amount of progeny was present in the culture bottle the P1 generation was killed.

After the progeny developed in the F1generation of the sex-linked cross the phenotypes were observed and recorded. After the F1 generation of the monohybrid cross developed the process was repeated to create an F2 generation and the results were observed and recorded. Class data was also included in the results for comparisons. We found that though some of the results for each of the crosses were close to the null hypothesis, they did not accept the probability of the Chi-Square analysis. This means that even though there are expectations for the results, what happens in nature can almost never be controlled completely.

Background

The first experiments utilizing the fruit fly Drosophila melanogaster were performed by Thomas Morgan, when in 1910 he spotted a white-eyed fruit fly in his fly room. However the first reported “mutants” found among the fruit fly were observed in 1907 by Frank Lutz of the Carnegie Institution. He reported different wing patterns and, by 1908, had noticed the proliferation of dwarf mutants. By the 1980s these discoveries have spawned into its own field of study by geneticists sometimes referred to as “Drosophilists”. (Taubes 2012)

The purpose of our D. melanogaster Monohybrid Cross experiment was to study the genetic traits based on Mendelian inheritance. In this cross, the wild type (red-eyed) phenotype is 100% dominant over the recessive sepia-eyed trait. The null hypothesis for the F1 generation is 100% wild-type and the F2 generation has a null hypothesis of 75% red-eyed and 25% sepia-eyed. (lab manual).

The purpose of our D. melanogaster Sex-Linked Inheritance experiment was to observe that not all traits are acquired autosomally. Through this lab we were able to see that X-linked traits are inherited in predictable patterns because of their dependence on the sex chromosome. The result of the crosses depends on which parent carries the phenotype wished to be observed before the actual cross. Since we crossed white-eyed females with red-eyed males the null hypothesis for this experiment was expected to be a 1:1 ratio of white-eyed males to red-eyed females (lab manual). Because males only carry one X chromosome (inherited from their mother which also determines eye color) they are always hemizygous for the sex chormosomes. (N.D. 2009).

Experimental Set-up:
Monohybrid Cross:
On August 28, 2012 a culture bottle with Drosophila media and yeast was prepared and labeled. Six red-eyed fruit flies (genotype RR) of one sex and six sepia-eyed fruit flies (genotype rr) of the other sex were transferred into the culture bottle after being anesthetized. After a period of about seven to ten days numerous larvae and some pupae became visible inside the bottle. To prevent any intergenerational mating, the P1 generation (original adult parent flies) were removed from the bottle leaving only the offspring to emerge into the F1 generation.

The F1 generation was apparent inside the culture bottle after another seven to ten day period. Using FlyNap these fruit flies were put under anesthesia for observation. The phenotype of the F1 generation fruit flies was observed to be all red-eyed; therefore the genotype was noted to be Rr.

On September 11, 2012 a new culture bottle with fresh media and yeast was prepared and labeled. Six male flies and six female flies from the F1 generation were added to the new culture bottle. Once larvae and pupae became visible the adult F1 generation fruit flies were removed from the culture leaving behind the F2 generation offspring. Seven to ten days later the offspring had developed into adult flies. The F2 generation was anesthetized and then separated and counted based on eye phenotype which was either red or sepia.

The numbers observed were used to calculate a Chi-Square statistical analysis which was used to determine whether or not the null hypothesis of a 3:1 ratio for the F2 generation was accepted. This data was combined with data from the entire class to increase the sample population and to calculate another Chi-Square.

Sex-linked Cross:
A culture bottle with Drosophila media and yeast was prepared and labeled. Six D. melanogaster wild-type male flies and six white-eyed female flies were transferred into the bottle. Seven to ten days later numerous larvae and some pupae became visible inside the bottle. At this time the P1 generation flies were removed from the bottle leaving only the F1 generation progeny as to avoid intergenerational mating.

Adult flies of the F1 generation were visible after a week. These adult flies were anesthetized and the phenotypes (sex and eye color) were observed and recorded. A Chi-Square statistical analysis was calculated to determine if the null hypothesis of 1:1 for fly sex and eye color would be accepted or rejected. This data was combined with data from the entire class to increase the sample population and a second Chi-Square was calculated.

Results
The P1 Generation for the monohybrid cross consisted of six homozygous red-eye (RR) fruit flies of one sex and six homozygous sepia (rr) fruit flies of the other sex. As shown in Table 1 the F1 pairing consisted of being all heterozygous for the wild-type red-eye (Rr) in coloration. The F2 generation is also presented showing an unexpected ratio and the Chi-square is calculated in Table 2. The P1 generation of the sex-linked inheritance started with white-eyed females and red-eyed males and the results in the F1 progeny are presented in Table 3 while the Chi-square is illustrated in Table 4.

Monohybrid Cross:
The total observed numbers counted was 358 with 333 (93%) of them having the red-eye phenotype and a genotype(s) of homozygous RR and/or heterozygous Rr. The remaining 25 (7%) were of the sepia-eye colored phenotype with a definite genotype of homozygous rr. The expected ratio of the null hypothesis was 3:1 (75%) red-eye colored and 1:3 (25%) sepia-eye colored or expected numbers of 268.5 (red) and 89.5 (sepia). P1 Eye Genotype &
The Chi-Square calculations were figured by taking the standard Chi-Square computations as shown in Table 2. The Chi-Square number for our experiment was 62.0. This number was significant in comparison to what would have been needed for the null hypothesis to have been accepted, thus enabling the conclusion of a rejected hypothesis. The class as a whole also calculated the Chi-Square by adding up all the observed numbers and expected numbers to form a class total. This figure was significantly larger at a Chi-Square of 206.7, also leading to a rejected null hypothesis.

Sex-Linked Cross:
The P1 genotype and phenotypes consisted of: XwXw white-eyed females and XwY Red-eye males. Six of each was crossed and the F1 generation was shown to have consisted of: Xw+Xw red-eye females and XwY white-eye males. The expected ratio was 1:1 red to white. With a total count of 357 fruit flies, the observed number of red-eye females was 220 while the observed number of white-eye males was 137 which did not fit the expected ratio.

The Chi-Square for our group was found to be 19.3 which was too high to be accepted, thus the null hypothesis was found to be rejected. The Chi-Square of the class was found to be 50.0 and once again too high to be accepted, thus the null hypothesis was rejected.

Conclusion
The results of both of these experiments were similar in terms of the acceptance or rejecting of the null hypothesis. The expected probability ratios were theoretically supposed to have closely mimicked the experimental results; however probability is just that, probable. There is no guarantee, due to randomness of the system, that there will be a ratio that will ever exactly mimic real life scenarios in nature.

In both experiments the Chi-Square figure showed that both null hypotheses were rejected. These numbers were not comparably close to those needed to have had an accepted null hypothesis.  The fertility of these fruit flies, the frequency of mating, the genes that were involved with crossing over, and the chromosomes that were passed on to the next generation(s) were all due to random assortment and therefore it could only be just a possibility that certain traits pass on to the next generations. Mutations can also influence such ratios with traits that were not accounted for in the past and are consequently passed on to future generations. (such as albinism).

References

Genetics Lab Manual. (2012).
N.D. (2009). Sex-Linked Inheritance: Drosophila. National Health Museum. Retrieved October 5, 2012 from: http://www.accessexcellence.org/RC/VL/GG/sex.php.
Taubes, G. A. (2012). The “Fly People” Make History. The Genes We Share. Retrieved from http://www.hhmi.org/genesweshare/b100.html.