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UV mutagenesis lab.doc

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Northeastern University
BIOL 2301

UV Mutagenesis Melissa Johnson Maggie McCrann Lab period: Tues 2:50 Submitted: 12/03/07 Johnson 2 Introduction The objective of this exercise was to design and carry out an experiment to test how an outside variable affects the capability of UV mutagenesis to prevent the growth of Serratia Marcescens (Begley 2007). Serratia Marcescens is a bacillus shaped bacteria discovered by Bartolomeo Bizio in 1819. In the 1960’s S. Marcescens was believed to be non-pathogenic and was used in school experiments to track infections. The objective of the commonly used “handshake test” was to demonstrate the importance of hand washing by having one person dip their hand in the bacteria and shake someone else’s hand, and that person would shake another person’s hand, etc. The microbial transmission of S. Marcescens was then easily tracked due to its blood- red color (Begley 2007). Also, in the 1950’s S. Marcescens was used by the U.S. Army in “Operation Sea- spray” in order to test how wind currents would affect the spread of biological weapons. To test this, balloons were filled with S. Marcescens and burst over San Francisco, however not long after; doctors noticed an increase in pneumonia and urinary tract infection in the area. S. Marcescens has since been classified as an opportunistic human pathogen. The red pigmentation of S. Marcescens is caused by the expression of the pigment prodigiosin. Treatment for infection is antibiotic therapy, however, many strains of the bacteria are resistant to multiple antibiotics because of the presence of R-factors which are plasmids that can carry genes for resistance (Begley 2007). Ultraviolet (UV) radiation is a type of radiation received by the earth in the light from the sun. Its wavelength is shorter than that of visible light, and therefore, more harmful. Its name was derived from the fact that its electromagnetic waves have Johnson 3 frequencies higher than those that can be seen by humans as the color violet (Hartl 2006). UV radiation is harmful to organisms because it causes mutation in DNA (Begley 2007). DNA absorbs UV radiation which changes the shape of the molecule, causing mutations. UV radiation passes easily through air, not as easily through liquids and poorly through solids. Therefore, the bacteria to be tested must be directly exposed to a UV light. The effects of radiation are easily seen in bacteria because due to the fact that they are single-celled organisms, UV light can have very damaging effects on their DNA. The effects of UV radiation on human tissues can include sunburn, retinal damage, wrinkles and cancer (Begley 2007). Materials and Methods A killing curve was first established in order to test the exposure time needed for UV light to kill the bacteria. Five plates were tested at different time intervals by exposing one half of the plates to UV light and covering the other half with an index card for comparison. The results of the killing curve were observed and recorded below and helped to establish the exposure times used in the experiment (Begley 2007). Two possible experiments were discussed. The first was to test how distance from the source of radiation would affect UV mutagenesis of the bacteria. The hypothesis would be that as the organism moved further away from the source of radiation, less UV mutagenesis would occur. This experiment would be set up by exposing plates to UV light from five different distances at 30 second and 2 minute time intervals and observing the UV killing that occurred. It was concluded that this experiment would involve too much error and the following experiment was used. Johnson 4 The experiment that was carried out was designed to test the how well different SPF levels of sunscreen protect bacteria from UV mutagenesis due to exposure to UV light. Four levels of SPF and zinc oxide were tested; SPF 4, SPF 15, SPF 30, SPF 45, and Zinc Oxide. Two plates were tested with each level of SPF; one plate was exposed for thirty seconds, and one was exposed for two minutes to UV light. A control was also exposed to UV light with just saran wrap over the bacteria at thirty seconds and two minutes. Each plate used was covered in saran wrap and the sunscreen being tested was spread over half the plate. This half of the plate was then exposed to radiation. The plates were left to incubate and the results of the growth were observed and recorded below. Results Killing Curve: Plate Degree of Killing 1 1 2 2 3 3 4 5 5 5 Plate 1: 15 seconds Degree of Killing: Plate 2: 30 seconds 1 – least amount of killing Plate 3: 1 minute 2 Plate 4: 2 minutes 3 Plate 5: 5 minutes 4 5 – total killing Johnson 5 K illingC urve 6 5 4 3 KillingCurve 2 1 0 0 50 100 150 200 250 300 350 Sunscreen: Plate 30 seconds 2 minutes A 20+ colonies 1 colony B Less dense lawn that at 2 Lawn on ¾ of plate, few minutes, more single single colonies colonies C Lawn Lawn D Lawn Lawn E Lawn Lawn F Lawn Lawn Plate A: Control, no SPF Plate B: SPF 4 Plate C: SPF 15 Plate D: SPF 30 Plate E: SPF 45 Plate F: Zinc Oxide The growth on plate A, the control with no sunscreen, was the most growth observed. This was expected as there was no sunscreen on this plate. Plate B had the SPF 4 sunscreen and it was observed that there was more growth on this plate than the Johnson 6 control plate which was expected, however there was still a lot of UV killing. Plates C-E al had lawns of growth at both times. The expected results would be that the growth would gradually increase with greater SPF and lower time, however, the actual observed results showed much less UV mutagenesis than expected and there was more growth than expected on plates C and D. Discussion and Conclusion This experiment was designed to verify that higher levels of SPF in sunscreen will protect better from UV damage. Therefore, as the SPF levels increase at both time intervals, the ability of UV radiation to inhibit microbial growth will be reduced. The results obtained in this experiment gave a broad look at how well sunscreens of different SPF can protect from UV damage. It was observed that the sunscreen with the lowest SP
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