PHAGE HUNTER BY TOSHA THOMPSON Frederick Twort Felix

PHAGE HUNTER BY: TOSHA THOMPSON

Frederick Twort Felix D’Herelle BACKGROUND INFORMATION http: //es. wikipedia. org/wiki/F%C 3%A 9 lix_d'Herelle http: //en. wikipedia. org/wiki/Frederick_Twort • Bacteriophages were discovered by Frederick Twort in 1915 an English bacteriologist and Felix D’Herelle in 1917 who was a French-Canadian microbiologist. • During WWII phages were used to treat bacterial diseases, among soldiers, like gangrene and dysentery. References: * http: //phages. org/

PHAGE SAMPLE SOURCE: • Dr. Klyczeks compost pile • Collected: 9/21/13 @ 9: 30 am • Location: • Partially degraded compost (kitchen scraps and yard waste) from pile on west side of garage in Hudson, WI • GPS coordinates: 44° 59’ 4”N, 92° 45’ 16” W • Soil condition: very moist with partially degraded plant material • Soil depth: 30 mm • Air temperature: 66°F • Weather condition: dry & sunny

Koethe 95 PHAGE NAME • KOETHE 95 • This is a mixture between my boyfriends last name and the year I was born. • I named my phage after him because he is an important person in my life and stands beside me when I make decisions. At the University of Minnesota we got to take pictures of our phages underneath an electron microscope.

UNIVERSITY OF MINNESOTA • November 16, 2013 our class was invited to the U of M to take pictures of our phages • To the right is a picture of my phage: Koethe 95 @ 100 times its actual size • He’s considered to be a Silphoviridae because of his long tail • Head: 18. 75 nm (nanometers) • Tail: 112. 50 nm Koethe 95 Above is the picture of my phage: Koethe 95 which was taken at the U of M at 100 times its actual size.

PLAQUE MORPHOLOGY Before… Now Small plaques: . 5 mm Big plaques: ≈1 mm Appearance: *Cloudy *No halo In the beginning my plaques started out as a mixture of small and large sizes, as seen on the left. But as I continued for about a month of testing I finally got uniformed plaques which was the larger plaques. This was surprising because it seemed as if the small plaques were more dominant. Titration This is the 10^0 plate after my first titration which was taken on October 3, 2013. High Titer Lysate (HTL) Titration This is a picture of uniformed plaques after the fourth titration which was taken on October 22, 2013.

DNA ISOLATION 1 2 3 4 5 6 7 • DNA concentration: • Enzymes . 1425 ug/ul • Yield: 1 • 14. 25 ug • A 260/280 ratio: • 1. 781 • High Titer Lysate (HTL): Lane 1: Kb ladder Lane 2: Undigested Enzyme Lane 3: Bam. HI Lane 4: Cla. I Lane 5: Eco. RI Lane 6: Hae. III Lane 7: Hind. III • 1. 08 X 10^8 Restriction Digest Results Above is the restriction enzyme digest results from the gel test we conducted. In each of the lanes, labeled 1 -7, we used different enzymes to differentiate the phages in our classroom. By doing so we saw if the cuts made by the enzymes were similar to that of anyone else's in the classroom. This helped us determine if students had the same phage.

SECOND DNA GEL • I didn’t have the chance to do my second DNA gel because we didn’t have enough enzymes any longer and when we tried to use different enzymes it didn’t cut the DNA. • Because of this it leads me to believe that my phage could possibly be similar to that of Ashley Bue’s phage: Gathje • But I don’t have enough evidence to prove that they are the same so as of right now they are different phages

PHAGES THAT MAY BE SIMILAR 1 2 3 4 5 6 7 Enzymes 1 2 3 4 5 6 7 As you can see our DNA was cut in similar spots by the same enzymes. Which leads me to think that I might have the same Phage as Ashley Bue’s. But sense I don’t have a second gel like she does I have nothing to compare hers to mine with. So I can not tell you for sure if they are the same phage or not but I can say there is a possibility that they are the same. Lane 1: Kb ladder Lane 2: Undigested Enzyme Lane 3: Bam. HI Lane 4: Cla. I Lane 5: Eco. RI Lane 6: Hae. III Lane 7: Hind. III Ashley Bue’s GEL Tosha Thompson’s GEL (Mine) Above are two pictures of different gels. On the left is Ashley Bue’s gel from her phage Gathje. To the right is a picture of my gel from my phage Koethe 95. This is showing a comparison of the two phages this helps us determine a little better whether or not they are similar. But because I haven't done a second Gel we can’t be certain of anything as this point.

KOETHE 95