Modes of Inheritance and Mutations in Drosophila melanogaster
Nikya Allen
Texas State University
BIO 2450 Genetics
October 15th, 2018
Izzy DeLeon
Abstract
In this study, we wanted to see how the modes of inheritance change in each mutation we had. We were observing Mendelian genetics and how it occurred throughout each generation of D. melanogaster. From this study, we were able to get a better understanding of how each mode of inheritance works and take that new understanding to understand human mode of inheritance as well. We composed four different hypothesis for each mutation, ranging from six-linked, autosomal, dominant, or recessive. We looked at three different generations of D. melanogaster from beginning of life cycle to the end each weeks and took note of the number of mutants and wild types. When the testing was over chi-square testing was preformed to determine the significance. Only one set of mutation proved to be significant with our hypothesis, which was the vestigial mutation. Through this study we got a good look at the different modes of inheritance in D. melanogaster.
Introduction
In this study we were out to seek how the genes and chromosomes were working in D. melanogaster. We wanted to find out if the mutations were sex linked or autosomal. Because the D. melanogaster are small and reproduce rapidly it was easy to follow many generations. Unlike in humans how we only produce a few offspring and the chance of being a male or female is 50:50 so we could produce 2 females and would need a lot more offspring in order to evaluate what we are trying to study. We are able to use the Drosophila though to determine how Mendelian genetics works and use it for the benefit of human research. Mendelian genetics basically proposes that genes come in pair and are inherited as separate chromosomes from each parent.
The Drosophila are easily distinguishable among sex, are also harmless, and don’t spread any disease. Because of their ability to be so easy to work with thousands of laboratories, including ours, has taken advantage of their ability to produce about 25 generations a year(Smith, J). The rate of development is depends of the temperature they are stored at. At 20 degrees Celsius the life cycle is completed in 14 or 15 days, while at 25 degrees the cycle is completed in 10 days(Jorden, S.L.). Because the D. melanogaster only have 4 pair of chromosomes, it gives labs the ability to map single mutations along their chromosomes. Chromosomes 2, 3, and 4 are autosomes while the first pair, 1, is sex chromosome. Just like humans the female is XX and the male has CY. At the Bloomington Drosophila Stock Center, they were able to map the female-sterile mutation sat^SC46 mapped to Orc1, and able to determine that it must’ve been a partial loss of function allele because the knockout alleles are recessive lethal(Kahsai).
Sex linked genes and autosomal genes are very differently passed on from parent to offspring and the show up quite differently throughout each generation. Autosomal-linked traits are genes that are on the autosomes; sex-linked traits are because of the genes located on the sex chromosome. Sex-linked inheritance was first discovered by T.H. Morgan by studying Drosophila(Ashburner, M). From this type of inheritance, we were able to see how reciprocal crosses were not identical, and how the genes were located on the X or Y chromosome or both XY chromosomes. Similar to Morgan’s study we systematically carried out a series of genetic crosses to explain the inheritance of each mutation observed.
In order to explain what the inheritance of each mutation we had to begin with test crosses to prove where the gene was located, either on the sex chromosome or autosomes and if it was dominant or recessive. It is easy to determine between autosomal dominant and recessive. In order to determine if a gene is sex-linked dominant or recessive, it is easier to look at the generations as they move down. In sex-linked recessive, males are effect a lot more than females. It is hard to determine sex-linked recessive in females because they have two X chromosomes, while the males only have one X chromosome so it’s a lot more prevalent in them than in females. If the trait is sex-linked dominant the condition is expressed in heterozygous females and well as the males. Because of this sex-linked dominant is relatively rare in comparison to sex-linked recessive mutations or diseases. In autosomal recessive genes, a gene alteration must be present in both sets of chromosomes in order to be expressed located on an autosome. Organisms with just one copy of the gene do not have the mutation. Males and females have the condition in roughly equal proportions . In autosomal dominant, an alteration in one copy of the gene is need to cause a mutation or disease to be expressed and it is located on an autosome. When autosomal dominant is present you usually see it present from generation to generation, depending on the parents homozygosity.
The purpose of our lab was to see what the mode of inheritance was for each mutation we had. For each mutation we had a different hypothesis based on where the mutation gene was. The very first mutation was white eyed and we hypothesized that to be sex linked recessive. If the mutation for white eyes was sex linked recessive then it would have a ratio of 1:1:1:1. The next mutation was ebony and we hypothesized that to be autosomal recessive. If ebony was autosomal recessive then we would see a ratio of 3:1, 1 being the mutation. The next mutation was vestigial and we hypothesized that to be autosomal dominant. If the ebony mutation was autosomal dominant then we would see a ratio of 3:1, with 3 being the mutants and 1 being the wild type. Lastly, we hypothesized the sepia eye mutation to be autosomal dominant. If the sepia mutation was autosomal dominant then we would see a 3:1 ratio, with 3 being the mutation and 1 being the wild type.
Materials and Methods
The parental generation Drosophila were held in a vial with a large amount of media and a vial plug. Once obtained, the tubing for CO2 to flow to the pad and blow gun was turned on. In order to ensure Drosophila weren’t stuck in the media once anesthetized, the vial was inverted. Making sure the tip of the blow gun wasn’t inserted past the plug, CO2 was slowly depressed into the vial. The Drosophila were then anesthetized and dropped down to the plug within a few seconds. They were carefully removed and placed onto the CO2 pad. Using a small paint brush, the Drosophila were sorted on the CO2 according to sex and presence of mutation. In order to see the Drosophila clearer the CO2 pad was placed under a stereoscopic microscope. Once the examination process was finished the Drosophila were placed back into the vial, by turning the vial horizontally and carefully brushing them into the vial. The flies were then placed back into the incubator set from 16-29 degrees Celsius. This same process was then repeated the following weeks with the F1 and F2 generations.
Results
In white eyed mutation we expected it to be sex linked recessive with ratio 1:1:1:1. In the ebony mutation we expect it to be autosomal so 3:1 ratio. In the vestigial mutation we expected it to be autosomal so 3:1 ratio. Lastly, the Sepia mutation was expected to be autosomal with a 3:1 ratio.
Table 1- Observed Phenotypes of Mutations
Mutation Wild Type Females Mutant Females Wild Type Males Mutant Males
A- White Eye 73 81 27 56
B- Ebony 40 40 45 18
C- vestigial 14 69 34 52
D- Sepia Eyes 44 41 15 24
D’- Sepia Eyes (my group alone) 28 39 8 22
Table 2:- P-Values and Chi Squared of Mutations
Mutations Ratio X^2 Degrees of Freedom P- Value
A- White Eyes 1:1:1:1 29 3 Less than .0001
B- Ebony 3:1 18 1 Less than .0001
C- Vestigial 3:1 1 1 .3173
D- Sepia Eyes 3:1 33.7 1 Less than .0001
D’- Sepia Eyes (my group alone) 3:1 7.60 1 .0058
Description for Table 2:
We fail to reject the null hypothesis of the mutation C, vestigial.
In each of the other mutations we fail for reject because the p-value was less than than 0.05.
In mutation A, white eye, we expected a 1:1:1:1 ratio because of the sex linked trait. The chi squared number was 29 with 3 degrees of freedom, and a p-value of less than .0001.
In mutation B, we expected a 3:1 ratio, the wild type being dominant and the mutation being recessive autosomal. The chi squared number was 18, with 1 degree of freedom, and a p-value of less than .0001.
In mutation C, vestigial, we expected to see a 3:1 ration with the mutation being autosomal dominant and the wild type being recessive. The chi squared number was 1 with 1 degree of freedom, and a p-value of .3173.
In both cases of mutation D, sepia eyes, we expected to see the ration 3:1, with the mutation of sepia eyes being the autosomal dominant trait and the wild type being recessive. In the combined sepia mutation the chi squared value was 33.7, with 1 degree of freedom, and a p-value of less than .0001. In just my group alone of the sepia eye mutation, the chi squared number was 7.60, with 1 degree of freedom, and a p-value of .0058. There was error between my group alone and when the two groups of sepia eye combined.
Discussion
When analyzing our results, we used the chi square testing. We determined what we thought each mode of inheritance was in our hypothesis. From each hypothesis there was a ratio that went with it and it had an expected amount of each organisms between male and female and mutant and wild type. From that we were able to preform a chi square testing that giving us a p-value letting us know if our data was significant or not significant. If the p-value is less than .05 than the data is nonsignificant, and we reject the null hypothesis. If the p-value is greater than .05 than the data is significant and we fail to reject the null hypothesis. When we fail to reject the null hypothesis that means you don’t have enough evidence to reject its statement or prove it wrong.
In our study, we started off with homozygous mutant females and homozygous wild type males for each mutation. With the white eyes mutation it was hypothesized that it was X chromosome sex linked, and the data proved to be not significant with our hypothesis, our p-value was less than .0001. We thought this was sex-linked recessive because it followed the 1:1:1:1 ratio closer than it did to the 3:1 ratio and more males were affected than females, which is known to occur frequently in sex-linked recessive genes. Next we analyzed the ebony mutation and hypothesized it to be autosomal dominant with a 3:1 ratio. Our studies were not concise with out hypothesis as our p-value was out of range and it was not significant, our p-value was less than .0001. The next mutation was Sepia eyes, and we predicted that to be autosomal recessive. The p-value was not significant with our hypothesis. As two groups combined our data was not significant because the p-value was less than .0001. With just my group alone, the data was actually significant, our p-value was .058, meaning that we fail to reject our null hypothesis. The only mutation that showed to match up with our hypothesis as a group all together, was the vestigial mutation. Because our p-value was greater than .05, .3173, meaning there is weak evidence against our null hypothesis, and we fail to reject it.
There were several human errors that occurred throughout our experiment. Some of the human errors included flies falling off the CO2 pad, accidentally killing some flies by handling them too rough, not placing the CO2 gun in the right position when anesthetizing the flies, and many more situations may have occurred in other groups. Another error that occurred when conducting our experience is that when we were finishing up counting the last generation of flies the other group in the sepia eye mutation didn’t separate the number of males and females and just divided the flies by mutation and wild type and then divided the males and females up evenly. That may have messed up with our data and could be the reason why just my group alone data was significant, meanwhile our numbers added together in total wasn’t significant. Another reason for error may have been stemmed from the fact we have a very small sample size, the larger the sample size the easier it is to see Mendelian genetics working in a population.
Mendel’s law states that hereditary units occur is pairs and separate for each gamete(Webster). We saw this occurring throughout our study from each mutation and seeing each mutation getting passed on and expressed differently in each offspring because of Mendel’s Law of segregation. Mendel’s second law basically states that the different pairs of units are distributed independently random to each gamete(Webster). This was shown in our study because it was very random and different from offspring to offspring. Mendel’s last law states that the genes will be expressed as dominant, unless both factors are recessive. This is shown throughout our study because none of our mutations were shown partially, it was either dominant or recessive.
For future experiments we could repeat the same experiment with a much larger sample size to see how that effects or data. We could also take flies with multiple mutations and see if they affect each other at all.
From our four different mutations we got go take a good look at how the mode of inheritance in D. melanogaster work. Even though most of our data wasn’t significant we got to see how the mutations were passed on from generation to generation and how it worked all the way from the parental generation to the F1 and F2 generations. From observing the phenotypes associated with certain mutations we were able to gain insight into the function of genes.
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