The Life Cycle of Drosophila Lab report

Monohybrid cross of wild type and wingless Drosophila melanogaster

Abstract

The principles of classical genetics are used to predict how genes determine basic patterns of inheritance, and are frequently explored using the common fruit fly, Drosophila melanogaster. Not only does such research provide basic information about inheritance, it can be appliced to human inheritance. In this study, pure bred cultures of wild type dominant flies were crossed with pure bred mutant apterous flies, producing both F1 and F2 generations. While the F1 generation was all of the wild type, the F2 generation produced a ratio of 3 wild type flies to 1 apterous type lies. This ratio supports Mendel's laws of inheritance. Using such research reinforces the value of understanding these principles, and also opens the need for future research regarding the conservation of genes in the human genome, which has a practical application in the study of human genetic diseases.

Introduction

Gregor Mendel, who is referred to as the father of genetics, used evidence he gathered through selective breeding of pea plants to establish the basic laws of genetics in the mid 1800's. These laws form the basis of classical genetics because they predict how genes behave using basic patterns of inheritance. Mendel developed these laws before genes had even been identified or understood, but the principles still hold true today. They are often studied in classical genetics experiments, such as is used with the common fruit fly, Drosophila melanogaster. This model organism is used because it is easily cultured, produces a new generation within two weeks, produces up to 500 eggs in ten days, is small enough to be studied in situations with limited space, but is individually large enough for easy phenotype determination (Flagg, 2005). Because the fly has been studied for over 100 years, many genes of D. melanogaster have been identified. This identification is useful because many biological properties are similar in mammals and D. melanogaster, and about 75% of the genes that cause diseases in humans are similar to functional genes in the fly (Pandey & Nichols, 2011). Furthermore, the sequencing of the D. melanogaster genome, which was first published in 2000, provided many contributions to genome research and genome projects(Ashburner & Bergman, 2005). In this experiment, D. melanogaster strains are crossed to study the basic laws of inheritance of the flies using the wild type and wingless, or apterous, strains. The experiment is designed to understand the basic principles of a monohybird cross, to learn how to cultivate D. melanogaster, and to explain how autosomal dominant and recessive genes are inherited, including how the alleles determine the phenotypic appearance of the flies. If the wingless phenotype of D. melanogaster is inherited according to Mendel's laws for a simple dominant/recessive autosomal gene, then the phenotypes of the F2 generation should produce a Mendelian ratio of 3 wild: 1 wingless.

Methods

Materials used in the experiment included culture vessels and plugs, Drosophila culture medium, stock cultures of of homozugous wild type D. melanogaster (+/+) and homozygous apterous D. melanogaster (ap/ap) flies, markers, anesthetic, brushes, and a magnifying lens. Using culture vessels and Drosophila culture medium, each stock culture was self crossed using five flies per sex in each of the two self-crossed cultures. This cross produced two culture tubes of parent (P) generation wild type and two culture tubes of P generation apterous flies. After removing any adult flies,virgin females were obtained for each cross by selecting them within 15 hours of emerging from the pupae. The P generations were then crossed using five male +/+ and five female ap/ap in each of two cultures, and also five female +/+ and five male ap/ap in each of two cultures. This yielded the first filial ( F1) generation. The flies were anesthetized, and the phenotypes of all flies of the F1 generation were noted. Next, virgin females were obtained for each F1 cross as in the previously described procedure. The F1 flies were crossed by mating five males and five females from each of the two cultures. The data from both pairs was combined and tested for the 3:1 phoentype ratio, using chi-square distribution statistics to see if the observed data matched the expected Mendelian ratios for autosomal inheritance.

Results

When the P1 cultures were crossed, one being pure wild type (+/+) flies and one being apterous (ap/ap) to produce the F1 generation, 20 flies were collected. All of these F1 offspring were observed to be have a winged, or wild type phenotype. For the F2 generation, 73 flies were observed. Of the 73, 55 had the wild type phenotype, and 18 were apterous, or wingless.

Using chi square statistical analysis of these figures for the F2 generation based on the Mendelian expected ratio of 3 wild type:1 wingless the following results were obtained as shown in Table 1.

Phenotype	Observed (O)	Expected (E)	(O-E)2	(O-E)2 /E
Wild type	55	54.75	0.0625	0.0011415
Apterous	18	18.25	0.0625	0.0034246
				X2 =.0045661

Table 1.Sum of (O-E)2 /E for all classes of D. melanogaster

Discussion

Because all 20 flies of the F1 that were observed were of the wild type and were heterozygous, it indicates that the wild type gene (+) is dominant over the apterous gene (ap). For the F2 generation, of the 73 flies observed 55 were of the wild type and could either be (+/+ ) or (+/ap) because this is the phenotype produced by the dominant gene. The apterous pehnotype had to be (ap/ap) because it was determined it was the recessive trait, and was observed in 18 individuals. Applying the chi square test to this data based on observed numbers and expected numbers which would be obtained in a Mendelian cross producing a 3:1 ratio, chi-square value was calculated to be .0045661, as shown in Table 1 of the results section. Based on the relationship of probability to the chi-square value, this shows the chi-square value places the probablity at 97.5% that the observed data meets the 3:1 ratio, as shown in Table 2. This shows that the deviation is expected very frequently, 97.5% of the time, so the hypothesis regarding the expected Mendelian ratio is true.

This data is supported by the background material provided for the experiment. According to Flaag (2005) , when winged flies that are homozugous are mated with homozygous apterous flies, the offspring of the F1 generation all will be +/ap and have wings. Furthermore, matings using the F1 flies produces an F2 generation that has a ratio of three winged offspring to one apterous offspring. These results are known to occur because the wild gene is dominant and the mutant gene is recessive.

D. melanogaster has been studied for over 100 years; as a result, many genes of D. melanogaster have been identified. This identification is useful because many biological properties are similar in mammals and D. melanogaster (Pandey & Nichols, 2011) due to these genes. The ap gene is no exception. A human protein identified as LHX2 produces ap mutant phenotypes in the same manner as the fly protein. Furthermore, both proteins are expressed in the nerve cords, eyes, olfactory organs, brain, and limbs of both flies and humans (Rincón-Limas et al., 1999). This shows that the expression of the proteins comes from the genes of a common ancestor, and emphasizes the importance of conducting research using D. melanogaster.

	P
df	0.995	0.975	0.9	0.5	0.1	0.05	0.025	0.01	0.005	df
1	.000	.000	0.016	0.455	2.706	3.841	5.024	6.635	7.879	1
2	0.010	0.051	0.211	1.386	4.605	5.991	7.378	9.210	10.597	2
3	0.072	0.216	0.584	2.366	6.251	7.815	9.348	11.345	12.838	3
4	0.207	0.484	1.064	3.357	7.779	9.488	11.143	13.277	14.860	4
5	0.412	0.831	1.610	4.351	9.236	11.070	12.832	15.086	16.750	5
6	0.676	1.237	2.204	5.348	10.645	12.592	14.449	16.812	18.548	6
7	0.989	1.690	2.833	6.346	12.017	14.067	16.013	18.475	20.278	7
8	1.344	2.180	3.490	7.344	13.362	15.507	17.535	20.090	21.955	8
9	1.735	2.700	4.168	8.343	14.684	16.919	19.023	21.666	23.589	9
10	2.156	3.247	4.865	9.342	15.987	18.307	20.483	23.209	25.188	10
11	2.603	3.816	5.578	10.341	17.275	19.675	21.920	24.725	26.757	11
12	3.074	4.404	6.304	11.340	18.549	21.026	23.337	26.217	28.300	12
13	3.565	5.009	7.042	12.340	19.812	22.362	24.736	27.688	29.819	13
14	4.075	5.629	7.790	13.339	21.064	23.685	26.119	29.141	31.319	14
15	4.601	6.262	8.547	14.339	22.307	24.996	27.488	30.578	32.801	15

Table 2. Critical value of the X2 distribution.

(Griffiths A.J.F, Miller J.H, Suzuki D.T, et al. ,2000).

References

Ashberner, M. and Bergman, C.M. 2005. Drosophila melanogaster: A case study of a model genomic sequence and its consequences. Genome Research. 15: 1661-1667.

Griffiths AJF, Miller JH, Suzuki DT, et al. 2000. An Introduction to Genetic Analysis. 7th edition. New York: W. H. Freeman.

Pandey, U.B. And Nichols, C.D. 2011. Human disease models in Drosophila melanogaster and the role of the fly in therapeutic drug discovery. Pharmacological reviews. 63(2): 411-436.

Rincón-Limas, D. E., Lu, C.-H., Canal, I., Calleja, M., Rodríguez-Esteban, C., Izpisúa-Belmonte, J. C., & Botas, J. 1999. Conservation of the expression and function of apterous orthologs in Drosophila and mammals.Proceedings of the National Academy of Sciences of the United States of America. 96(5): 2165–2170.