Poppy

By: Sam Greeley and Temmy Lawson

Background Information

• The reason we are studying Phages is because they can be used in the medical field as antibiotics and there is not a lot of information about all the different types. We are helping by researching new phages and hopefully discovering them.

Name and Reason

•Poppy - We decided on the name Poppy because we thought it was cute and our phage POPS in the plates.

Phage sample source

• Date: September 08, 2013• Location: “My moms garden, close to a wooded

area, surrounded by dry grass and exposed to nature.” Breanna Krohn

• Soil condition: Dry and loose• Air temp: 66 degrees• Weather condition: Cloudy and windy• Other info: “Vegetables aren't growing too well,

wanted to test soil.”

Plaque MorphologySize: 4mmAppearance: Clear and includes some size variation. Size variation is constant, so we believe it is part of the DNA.

Titration #2

This is the image that we feel best shows our plaque colonies while we were still picking and processing them for DNA.

Titer of HTL (High Titer Lysate)

• Pfu/mL = (pfu/# µL)• 1000µL/1 mL • dilution factor• Pfu/mL = 68pfu/ 50µL • 1000µL/1mL

• 10-4 → 10,000• Pfu/mL = 13,600,000 → 1.36 x 107

DNA Isolation

• Concentration (µg/µL)A260 x 50 x dilution x 1mL/1000µL0.259 x 50 x 10 x 1mL/1000 µL = 0.1295 µg/µL• Yield (µg)

yield = µg/µL x total µL = µg0.1295 x 100 = 12.95 µg

DNA Isolation Cont.

• A260 to A280

- Ratio is 1.786 (about 2)- Interpretation

The A260 is where the DNA is absorbed the most and A280 is where proteins are absorbed the most. The A280 should be about half of the A260 so you know there is not protein contamination so ours is good to go!

DNA Isolation Cont.

Restriction Digest Results

Kb LadderBam HIClaIEco RIHaeIIIHindIIIUndigested DNAThis picture shows our results of electrophoresis. The first well, a kb ladder, is a reference well and the other five are enzymes. The last one is just our DNA. This tells us that the HindIII is the only enzyme that cut our plaque DNA.

DNA Isolation Cont. • We did not have enough DNA to sequence and

run a second gel. Unfortunately this means that we do not have another picture for analysis of our phage.

Electron Microscopy

• Head: 50 nm• Tail: 93.75 nm

This is a cropped photo of our original picture taken from the electron microscope at the U of M so that a more clear image of our phage could be seen up close.

Poppy

Analysis

Poppy Sueme

This is our phage versus Bre and Bri’s phage, Sueme, electron microscopy pictures. At first when we were analyzing our phages through titrations, streak plates and electrophoresis we thought that we had the same phages. After looking at these pictures and measuring the heads and tails, we think that our phages have similarities but are not the same. They may end up being the same but we will not be sure unless they are DNA sequenced.

Why pick Poppy?

• Poppy is awesome• It came from a vegetable garden, and has the

potential to be ingested by a lot of people• The original sample came from Somerset,

Wisconsin so it has a better chance of being a new phage not in River Falls.