Phage Status ReportEmily Lehmann and Ellie Kriese

Phages 101• Phage is short for Bacteriophage• The name Bacteriophage means, “eater of the bacteria”• Phages are viruses that infect bacteria• Benefits from discovery:

• Super Virus• Medical Therapy – use as a treatment for infectious diseases• Genetic Diversity – learn more about genetic information

All information came from Science News online magazine article, All the World’s a Phage written by John Travis.

BarretLemonEmily:

Lemon: In high school my friends would always call me Lemon from

my last name Lehmann.

Ellie:

Barrett: Named after my 7th grade biology teacher who inspired me

to pursue a degree in biology.

- Collected on September 22, 2013 at 3:30pm near Chippewa Falls Wisconsin

-GPS Coordinates: 41 degrees 54’ 35’ N 91 degrees 22’ 45” W

-Garden Soil that was located near tomato plants and under grass clippings, as well as compost which included food scraps and horse feces

-Soil was very loose and moist

Sample Source

• Plaque Size: Varies from 1mm to 3mm

• Clear without halo

• When we first started this experiment Ellie picked and went forth with the larger plaques and Emily picked and went forth with the smaller plaques.

Plaque Morphology

First Soil Sample Titer10^-4 plate

Ellie picked

3 & 4

Emily picked

1 & 2

Titer of HTL

43 Plaques on the 10^-6 plate

43plaques/10mL x 100mL/1mL x 10^6 =

4,300,000,000 PFU/mL

4.3X10^9 PFU/mL

DNA Isolation

Concentration: .135mg/mL

Yield: 13.5mg

A260/280 Ratio: 1.686

Which is slightly below the optimal amount of 1.8, meaning that there may be small amounts of proteins

and other debris in the sample.

Digest 1:

1 KB LadderUndigestedBamHIClaIEcoRIHaeIIIHindIII

Restriction Digest

Results: Unfortunately, none of the enzymes appeared to cut the DNA except minor cuts in HindIII, which is not enough to distinguish whether or not this is a unique phage.

Restriction Digest Digest 2:

NcoI

EcoRV

SalI

PstI

1 KB Ladder

Results:Again these enzymes did not cut the DNA again making it difficult to distinguish whether or not this is a unique phage.

Similarities to the Database

After looking at our Gel results:

- We felt the phage, Laryoe, was the most similar to our phage’s digest results

- For the enzymes, EcoR I, BamH I, and Cala I, both our phage and the data base phage showed little or no digestion with these enzymes.

Phage 1:

Head diameter: 1cm= 62.5 nmTail Length: 2.3cm= 143.75 nm

Phage 2:

Head diameter: 0.9cm= 56.25nmTail Length: 2.3cm= 143.75nm

Electron Microscope

12

Emily’s Electron Microscope Image

Phage 1: Head Diameter: 1.1cm= 68.78nmTail Length: 2.3 cm= 143.75nm

Electron Microscope

Ellie’s Electron Microscope Image

11

Conclusions

When analyzing the electron microscope images we were able to conclude that we have the same phage due to the length and

appearance of the tail. The head diameters were also fairly similar. The structure of our phage appears to be siphoviridae structure with a

very wide tail, making it unique from any other phage found in the class and database.

Why Choose our Phage-This phage was found in Chippewa Falls WI which differentiates it from the other phages many of which were found by Dr. Klyzack.

-There is enough DNA that was collected to send it for sequencing

-This phage is unique because while there was a variation in size of plaques, the appearance of the plaque did remain uniform.

-Also this phage continuously plagued out plates all the way through 10^-3, it produced many plaques when it was not diluted.