West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
1
Mutagenic Effects of Benzalkonium Chlorid...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
2
Table of Contents
Contents Page Number
In...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
3
Observation of Visible Mutations
Data Ana...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
4
Chapter 1
INTRODUCTION TO THE STUDY
Backg...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
5
In this experiment, benzalkonium chloride...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
6
Statement of the Problem
Generally this s...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
7
Significance of the Study
This study woul...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
8
Chapter 2
REVIEW OF RELATED LITERATURE
Be...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
9
is harmful as it may result to systemic e...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
10
and MN formation in mammalian and plant-...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
11
CHAPTER III
Research Design and Methodol...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
12
Preliminary Activities
Culture Media Pre...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
13
Preparation of culture medium with mutag...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
14
generation will again have their phenoty...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
15
Data collection and Analysis
All of the ...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
16
Chapter 4
Results and Discussions
This s...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
17
Graph 1: Fruitflies in the First Filial ...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
18
Table 2: Incidence of Mutation in the Fi...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
19
The results of our F1 generation showed ...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
20
The results in our F1 generation showed ...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
21
Graph3: Fruit Flies in the Second Filial...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
22
Table 5: Incidence of Mutation in the Se...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
23
The results of our F2 generation showed ...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
24
(Richardt, 2002). The spatial localizati...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
25
very different size and number; if only ...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
26
Chapter 5
Conclusions:
Based on the gath...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
27
Bundesinstitut fur Risikobewertung. (201...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
28
cells at environmentally relevant concen...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
29
glial cells. Retrieved January 13, 2012 ...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
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Appendi
West Visayas State University
College of Arts and Sciences
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31
Normal Wild-type Fruit Flies
Yellow-bodi...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
32
Ebony Fruit flies with blistered wings
E...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
33
Hunchbacked Ebony Fruit flies
Short-wing...
West Visayas State University
College of Arts and Sciences
La Paz, Iloilo City
34
Fruit flies with weird antennae
Fruit fl...
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Developmental Biology Special Problem: Benzalkonium Chloride and Drosophila melanogaster (Fruit Fly) findings

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Mutagenic effects of Benzalkonium chloride on the development of Drosophila melanogaster

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Developmental Biology Special Problem: Benzalkonium Chloride and Drosophila melanogaster (Fruit Fly) findings

  1. 1. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 1 Mutagenic Effects of Benzalkonium Chloride on Drosophila melanogaster In Partial Fulfillment of the Requirements In Bio 206: Development Biology By Christopher Jon I. Gaw Put Mary Grace C. Dumdum Joanna Rose B. Navarro BSBS 3B January 2013
  2. 2. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 2 Table of Contents Contents Page Number Introduction to the Study Research Paradigm Statement of the Problem Hypothesis Significance of the Study Delimitation of the Study Review of Related Literature Methodology Research Design Methodology Materials Chemicals Preliminary Activities Culture Media Preparation Collecting of Samples Larva of Drosophila Melanogaster Experiment Proper Preparation of Mutagen Induction of Mutagen Separation and Transferring of Fruit Flies
  3. 3. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 3 Observation of Visible Mutations Data Analysis Results and Discussions Conclusions References Appendix
  4. 4. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 4 Chapter 1 INTRODUCTION TO THE STUDY Background of the Study Without genetic variations, some of the basic mechanisms of evolutionary change cannot operate. Mutations, one of the primary sources of genetic variation, are the changes in the DNA. A single mutation can have a large effect. The fruit fly Drosophila melanogaster is chosen as the organism of choice because it is a favored organism for the study of genetic basis of development. It is used extensively in genetic breeding experiments. It is an ideal testing organism because it has a short life cycle, exhibits great variability in inherited characteristics and can be easily raised to produce large numbers of offspring (Sandhyarani,2011). Frequency of mutation in fruit flies has been well documented. Mutation, as an event that gives rise to heritable alteration in the genotype, in the fruit flies may be classified as wildtype or a mutant. Visible mutations are easily identified as the loss of a function or difference in the color of the wildtype fruit fly.
  5. 5. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 5 In this experiment, benzalkonium chloride was used as an inducer of mutation. It is an antiseptic agent, commonly found in household products used for cleaning. Previous studies have showed that benzalkonium chloride is a mutagen as it affects mammalian somatic cells, yeasts, or bacteria (ScienceLab.com) This study aims to determine if the 10% concentration of benzalkonium chloride affects the phenotype of Drosophila melanogaster. If mutation does occur, this study aims to observe the different mutant phenotypes and compare phenotypes between generations. Research Paradigm Independent Variable Dependent Variable Figure 1.Shows the relationship between the independent and dependent variables of the study Phenotype of Drosophila melanogaster Treatment A 10% concentration of Benzalkonium chloride
  6. 6. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 6 Statement of the Problem Generally this study will be conducted to determine the effect of the 10% concentration of benzalkonium chloride to the phenotype of Drosophila melanogaster. Specifically this study shall seek to answer the following questions: 1. What is the effect of the 10% concentration of Benzalkonium Chloride on the phenotype of Drosophila melanogaster? Hypothesis This hypothesis will be tested on this study. 1. Benzalkonium chloride does not significantly affect the phenotype of Drosophila melanogaster. Definition of terms
  7. 7. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 7 Significance of the Study This study would benefit the following: General public. This study will affirm whether benzalkonium chloride is indeed a mutagen. This will give information to people that it is a mutagen and they will be aware of its possible effects. This study can also benefit researchers as this can be a ground for this study. Delimitation of the Study This study is limited to assessing the mutagenic effects of benzalkonium chloride only.Only one concentration will be used for this study by solving its weight by volume to get its 10% concentration. There will be 1 trial for this experiment. Pineapple will be used to collect the fruit flies, and the offspring of the collected fruit flies will constitute the parental generation. Sweet potato will then be used as a medium during the actual conduct of the experiment.
  8. 8. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 8 Chapter 2 REVIEW OF RELATED LITERATURE Benzalkonium chloride is considered a quaternary ammonium compound (QACs) which are cationic surfactants that are widely used as disinfectants (BfR, 2012). In household products, they are primarily found in fabric softeners, detergents and hair products (Ferk, 2007). According to ScienceLab.com, benzalkonium chloride (BAC) is mutagenic for mammalian somatic cells, bacteria or yeasts. It contains a material which may cause damage to the kidneys, liver, heart, gastrointestinal tract, cardiovascular system, and central nervous system. Benzalkonium chloride (BAC)may affect genetic material and cause adverse reproductive effects based on laboratory experiments done on animals. Benzalkonium chloride may be harmful when ingested accidentally. Animal experiments have indicated that ingestion of less than 150 gram may be fatal or may produce serious damage to the health of an individual. It can also produce chemical burns to the eye and skin following direct contact. Skin contact
  9. 9. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 9 is harmful as it may result to systemic effects following absorption. Benzalkonium chloride is an antiseptic agent which must not be applied to areas which have not been fully rinsed as it is inactivated by organic compounds. Application of BAC may include disinfecting instruments and preserving drugs in low concentration form (chemicalland). In a study done by Ferk and his team, he studied the genotoxic effects of benzalkonium chloride (BAC) in mammalian and plant cells at environmentally relevant concentrations. BAC was tested in four genotoxicity tests in the Salmonella/microsome assay with strains TA 98, TA 100 and TA 102, in the single cell gel electrophoresis (SCGE) assay with primary rat hepatocytes and in micronucleus (MN) assays with peripheral human lymphocytes and with root tip cells of Viciafaba. Results show that with BAC, a moderate but significant effect was found at an exposure of 1.0mg/l. Findings show that this chemical includes moderate but significant genotoxic effects in eukaryotic cells at concentrations which are found in wastewaters and indicate that their release into the environment may cause genetic damage in exposed organisms. The results show that BAC that was tested cause DNA migration
  10. 10. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 10 and MN formation in mammalian and plant-derived cells but no gene mutation in bacteria. A disinfectant containing benzalkonium chloride has been identified as the most probable cause of birth defects and fertility problems in caged mice. In the cosmetic safety database, there is moderate to strong evidence that it is an immune, skin, and respiratory toxicant. One or more in vitro tests on mammalian cells exposed to BAC show positive mutation effects. According to an article by ET CBC News, scientists at the University of Ireland in Galway found that when they added the disinfectant benzalkonium chloride to common bacteria called Pseudomonas aeruginosa, increasing the amount of germ-fighting solution over time, the bacteria learned to survive. The researchers discovered that over time the bacteria became more efficient at ridding their cells of both the disinfectant and antibiotic. They also found that a mutation in their DNA made them resistant to antibiotics.
  11. 11. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 11 CHAPTER III Research Design and Methodology Research Design An experimental method was used in the study to assess the mutagenic effects of 10% benzalkonium chloride to Drosophila melanogaster. It consists of a 10% benzalkonium chloride concentration. Methodology Materials The materials that were utilized are overripe pineapple, sweet potato, wide-mouth bottles, cheesecloth, rubber bands, tray, medicine dropper, gas range, pan, fork/spoon, peeler, water, inoculating loops, cotton, ether/chloroform, dissecting and compound microscopes, camera, high precision balance and wild-type fruit flies. Chemical(s) The chemical that was used to anaesthetize the Drosophila melanogaster was chloroform.
  12. 12. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 12 Preliminary Activities Culture Media Preparation Overripe pineapple (Ananascomosus (l.) merr.) was used as the culture medium for this experiment. The medium was crushed until it is soft and placed in a sterile bottle to ensure a “no- mold” culture media. This was also be used for the collection of fruit flies prior to the experiment. Collecting of Samples The bottle with the cultured medium was placed beside a trash bin and was left there for several hours until the bottle was filled with fruit flies. The cheesecloth was then sterilized and was used to cover the sanitized bottle which was secured by the rubber bands. The collected fruit flies were left to breed until visible larva could be seen. Larva of Drosophila melanogaster The larva of the collected fruit flies was used as the parental generation when it reached its adult stage after six (6) days.
  13. 13. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 13 Preparation of culture medium with mutagen concentration: In preparing the 10 grams, 10% benzalkonium chloride concentration, the researchers weighed 1 mL of benzalkonium chloride via graduated cylinder which was followed by the addition of 10 grams mashed sweet potato. The mutagen and the culture medium were mixed thoroughly and were placed in a wide- mouth bottle. Culturing fruit flies and induction of mutagen: From the earlier collection of wild type fruit flies from the overripe pineapple, it was cultured using the sweet potato medium with benzalkonium chloride culture media. The fruit flies were allowed to mate and their offspring/s were perceived from larva to adult. Before the offspring develop into adult, the collected fruit flies were released so that only the parental generation will remain inside the bottle. The parents were then allowed to mate until they produced offspring which were our (F1) generation. When the offspring developed into adult, the females were separated from the males to prevent them from mating. Their phenotypes were observed and were also recorded. They were combined later to mate and their offspring (F2)
  14. 14. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 14 generation will again have their phenotypes be observed and recorded. Separation of Male and Female Virgins Drosophila melanogaster and Pairing and Transferring of Male and Virgin Female Drosophila melanogaster to a vial: The Drosophila melanogaster that was anesthetized were transferred in a microscope slide for clear viewing of the structures of male and female under a microscope. After viewing, the male and female virgin flies were then paired and transferred to a vial that contained the cultured medium. Observation of Visible Mutations The adult Drosophila melanogaster of the F1 generation and F2 generation were used in observing the visible mutations under the aid of a microscope. These mutations were identified based on the differences in appearance of a wild type fruit fly and a mutant one.
  15. 15. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 15 Data collection and Analysis All of the recorded data, the observations in the phenotype were collected for analysis. All the results will be presented in tabular form and graphs.
  16. 16. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 16 Chapter 4 Results and Discussions This study was conducted in order to assess the mutagenic effects of benzalkonium chloride on Drosophila melanogaster and to observe and compare subsequent mutations between generations. Parental Generation: 8 virgin females 5 males Table 1: Fruitflies in the First Filial Generation (F1 Generation) Fruitfly Number of Males: Male Frequency: Percentage: Number of Females: Female Frequency: Percentage: Wildtype 4 0.2 20% 7 0.35 35% Mutant 0 0 0 9 0.45 45% Total number: 4 16
  17. 17. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 17 Graph 1: Fruitflies in the First Filial Generation (F1 Generation) Our F1 generation showed us that out of the 20 fruit flies, there were 11 wild type fruit flies and 9 mutant female fruit flies. Out of the 9 mutant female fruit flies, there were 2 fruit flies that showed 2 kinds of mutation. The following graph and table will show you the incidence of mutation for the F1 generation in terms of its body color and wing shape. Wildtype Male 20% Mutant Female 45% Wildtype Female 35% F1 Generation
  18. 18. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 18 Table 2: Incidence of Mutation in the First Filial Generation in terms of body color(F1 Generation) Graph 2: Incidence of Mutation in the First Filial Generation in terms of body color(F1 Generation) Tan colored body (Male) 20% Tan colored body (Female) 35% Yellow colored body (Female) 35% Ebony colored body (Female) 10% Incidence of Mutations in the body of Fruit Flies (F1 Generation) Body Number of Males: Male Frequency: Percentage: Number of Females: Female Frequency: Percentage: Tan 4 0.2 20% 7 0.35 35% Yellow 0 0 0 7 0.35 35% Ebony 0 0 0 2 0.1 10%
  19. 19. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 19 The results of our F1 generation showed us that in terms of body color, it was found that only females expressed either of two mutant forms in this generation. Out of the total 20 fruit flies from the F1 generation, 4 males and 7 females expressed their tan coloration (wild type fruit fly). While there were, 9 females who showed either yellow or ebony coloration. Out of the 9 mutant fruit flies, 7 expressed the color yellow in their body and 2 expressed the ebony color. Table 3: Incidence of Mutation in the First Filial Generation (F1 Generation) Parts Number of Males: Male Frequency: Percentage: Number of Females: Female Frequency: Percentage: Short winged 0 0 0 1 0.05 5% Blistered wing 0 0 0 1 0.05 5%
  20. 20. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 20 The results in our F1 generation showed us that in terms of wing type and head type, it was found that out of the 9 mutant female fruit flies, 2 of which showed two kinds of mutation in terms of its body color and wing type. 1 fruit fly expressed a short type kind of wing and another 1 expressed a blistered kind of wing type. Table 4: Fruit Flies in the Second Filial Generation (F2 Generation) Fruitfly Number of Males: Male Frequency: Percentage: Number of Females: Female Frequency: Percentage: Wildtype 4 0.09 9% 13 0.32 32% Mutant 8 0.195 19% 16 0.390 39% Total number: 12 29
  21. 21. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 21 Graph3: Fruit Flies in the Second Filial Generation (F2 Generation) Our F2 generation showed us that out of the 41 fruit flies, there were 17 wild type fruit flies and 24 mutant fruit flies. Out of the 24 mutant fruit flies, there were 5 fruit flies that showed 2 kinds of mutation. The following graph and table will show you the incidence of mutation for the F2 generation in terms of its body color and wing shape. Wildtype Male 10% Mutant Male 19% Wildtype Female 32% Mutant Female 39% F2 Generation
  22. 22. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 22 Table 5: Incidence of Mutation in the Second Filial Generation (F2 Generation) Graph 4: Incidence of Mutation in the Second Filial Generation (F2 Generation) Ebony colored body (Female) 10% Tan colored body (Female) 32% Ebony colored body (Male) 19% Yellow colored body (Female) 29% Tan colored body (Male) 10% F2 Generation Body Number of Males: Male Frequency: Percentage: Number of Females: Female Frequency: Percentage: Tan 4 0.09 9% 13 0.317 31.7% Yellow 0 0 0 12 0.29 29% Ebony 8 0.195 19.5% 4 0.09 9%
  23. 23. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 23 The results of our F2 generation showed us that in terms of body color, it was found that out of the total 41 fruit flies, 24 mutant fruit flies expressed either yellow or ebony body coloration. Out of 24 mutant fruit flies, 8 males and 4 females expressed the ebony coloration. While there were 12 females that expressed the yellow body coloration. Discussions: Abnormal body coloration occurred most likely due to benzalkonium chloride affecting genes (the yellow genes), located on the X chromosome, which blocks the synthesis of brown pigment resulting to fruit flies with yellow bodies. Also, according to Bruce Alberts (1985), the yellow gene (y) is involved in pattern-specific melanin pigmentation of the cuticle of the adult fly and of larval mouth parts of Drosophila melanogaster and has mainly neural functions. This means that BAC is strong enough to alter such gene for the phenotypic observation of the mutant fruit flies that exhibited yellow body coloration. Other abnormal body coloration is due to a defect in their ebony gene which is responsible for the building up of the tan colored pigments in the normal fruit fly. Ebony was found to be exclusively localized to neuropile and epithelial glial cells
  24. 24. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 24 (Richardt, 2002). The spatial localization of Ebony protein within the adult nervous system, including the visual system and protocerebrum, was studied in order to examine the abundance of the protein at different times of day.Since the ebony gene is defective, this resulted to the black pigments accumulating all over the body thus we have obtained an ebony colored body fruit fly. In addition, according to Wittkopp, True, and Caroll (2002) Ebony is required to suppress some melanin formation, and is expressed in cells that will produce both melanizedand non- melanized cuticle. Ectopic expression of Ebony inhibits melanin formation, but increasing Yellow expression can overcome this effect. Another mutation that we have observed is with the wing type of the fruit flies. The short winged fruit flies have a defect in their vestigial gene on its second chromosome. Flies with short wing type have a recessive mutation (The Exploratorium, 2013). Short wing gene is located in chromosome I between 63.5 and 64.This gene proved to have a very pleiotropic manifestation, on changes in the wings and the eyes being the most common ones. The alterations in the wings are manifold; the wings may be stretched out in an abnormal direction, not backwards as is normal, but held out from the sides of the body at an angle of 90 degrees. The wings may exhibit incisions of
  25. 25. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 25 very different size and number; if only one incision is present this is usually found in the medial margin of the wing, but the medial, lateral and posterior margin may all have incisions (Eker, 2002). Blistered wings were also observed on the mutant ebony Drosophila melanogaster. According to Martin, Zusman, and Li (1999), a new laminin α chain is assigned to the locus of wing blister which is essential for embryonic viability and is involved in processes requiring cell migration and cell adhesion.Blistered gene in accordance to Fristrom, Gotwalset. al. (1994) both developmental and genetic analyses suggest that it is required during metamorphosis for the initiation of intervein development and the concomitant inhibition of vein development. A single wingless and legless Drosophila melanogaster was perceived on the second filial generation. Wingless trait according to Bishop, Huggler, et. al. (2010) or (mutation H) is an autosomal recessive gene. Meaning, two copies of an abnormal gene must be present in order for the disease or trait to develop.A mutation in a gene on one of the first 22 nonsex chromosomes leads to wingless trait.
  26. 26. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 26 Chapter 5 Conclusions: Based on the gathered results, the following conclusions were made: The results of the experiment showed that the Benzalkonium Chloride induces mutation to the F1 generation and F2 Generation of Drosophila melanogaster. The observed mutations in this study were: the yellow and ebony colored body of the fruit fly, and the short winged and blistered wing type. References: Alberts B. (1985). Molecular analysis of the yellow gene (y) region of Drosophila melanogaster.Retrieved January 14, 2013 from http://www.ncbi.nlm.nih.gov/pmc/articles/PMC391346/pdf/pnas 00361-0222.pdf
  27. 27. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 27 Bundesinstitut fur Risikobewertung. (2012). Health assessment of benzalkonium chloride residues in food. Retrieved on December 29, 2012 from: http://www.bfr.bund.de/cm/349/health-assessment-of- benzalkonium-chloride-residues-in-food.pdf Chemical land (2012).General Description of antiseptic agent. Retrieved on December 29, 2012 from: http://chemicalland21.com/lifescience/phar/benzalkonium%20c hloride.htm Chemwatch (2008).Benzalkonium chloride. Retrieved on December 29, 2012 from: http://datasheets.scbt.com/sc-257126.pdf Eker R. (2002).The Short wing gene in Drosophila melanogaster and the effect of temperature on its manifestation.Retrieved January 14, 2013 from http://www.ias.ac.in/jarch/jgenet/30/357.pdf ET CBC News. (2009). Disinfectant use could cause superbugs. Retrieved on December 30, 2012 from: http://www.cbc.ca/news/health/story/2009/12/28/disinfectant s-superbugs.html Ferk F. (2007). Benzalkonium chloride and dimethyldioctadecyl- ammonium bromide (DDAB), two common quaternanry ammonium compounds, cause genotoxic effects in mammalian and plant
  28. 28. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 28 cells at environmentally relevant concentrations. Retrieved on December 29, 2012 from: http://mutage.oxfordjournals.org/content/22/6/363.full.pdf Fristrom D., Gotwals P., et. al. (1994).Blistered: a gene required for vein/intervein formation in wings of Drosophila. Retrieved on January 14, 2013 from http://dev.biologists.org/content/120/9/2661.full.pdf Huggler A., Patel G. (2010). Analysis of the Mode of Inheritance of the Mutant White Eye Trait and Wingless Trait in live Drosophila melanogaster and the Mapping of Sepia Eye Color Mutation to Identify the Particular Chromosome on which the Mutation is Located in Drosophila using Virtual FlyLab. Retrieved on January 14, 2013 from http://by423-01-fall- 2010.wikispaces.com/BY423-01+Group+1+Report Martin D., Zusman S., and Li X. (1999).Wing blister, A New Drosophila Laminin α Chain Required for Cell Adhesion and Migration during Embryonic and Imaginal Development. Retrieved January 14, 2013 from http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2148222/ Richardt, A., Rybak, J., et al. (2002). Ebony protein in the Drosophila nervous system: optic neuropile expression in
  29. 29. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 29 glial cells. Retrieved January 13, 2012 from http://www.sdbonline.org/fly/sturtevant/ebony4.htm ScienceLab.com (2012).Benzalkonium chloride MSDS. Retrieved on December 30, 2012 from: http://www/sciencelab.com/msds.php?msdsId=9923038 The Exploratorium © (2013). Mutant fruit flies. Retrieved December 30, 2012 from http://www.exploratorium.edu/exhibits/mutant_flies/mutant_f lies.html WittkoppPJ., True JR., Carroll SB. (2002). Reciprocal functions of the Drosophila Yellow and Ebony proteins in the development and evolution of pigment patterns.Retrieved January 14, 2013 from http://www.ncbi.nlm.nih.gov/pubmed/11934851
  30. 30. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 30 Appendi
  31. 31. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 31 Normal Wild-type Fruit Flies Yellow-bodied Fruit flies
  32. 32. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 32 Ebony Fruit flies with blistered wings Ebony Fruit flies with proportioned body
  33. 33. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 33 Hunchbacked Ebony Fruit flies Short-winged Fruit fly Wingless and legless Fruit Fly
  34. 34. West Visayas State University College of Arts and Sciences La Paz, Iloilo City 34 Fruit flies with weird antennae Fruit flies with acute abdomen Solving for the 10% Benzalkonium Chloride Concentration: Mass/ Volume = 10g x 0.1 = 1 mL of Benzalkonium Chloride mixed with 9 mL of distilled water

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