Bacteriophage

Question 1

what was the purpose of this lab?

Answer 1

determine the concentration of T4r bacteriophage in a stock solution by carrying out a Plaque Assay

Question 2

what E coli are we using?

Answer 2

E coli B

Question 3

what two bacteriophages infect the Ecoli B

Answer 3

bacteriophage T4r and T2

Question 4

describe how the bacteria and phage will be immobilized in the top agar

Answer 4

a solution of dilute agar medium (top agar) containing a mixture of E coli B and phage is spread into tryptone agar (bottom agar) in a petri dish

Question 5

T or F: when a bacterial cell is infected by a phage, the bacterium will produce more viruses and eventually lyse

Answer 5

true

Question 6

when an infected bacterial cell lyses, what happens?

Answer 6

adjacent bacterial cells are infected

Question 7

describe a plaque

Answer 7

after several cycles of infection and lyses, a small area surrounding the point of infection that’s free of bacteria will be produced = a plaque

Question 8

how can the number of phages in the mixture be determined?

Answer 8

by counting the number of plaques, assuming each one is the result of one phage

Question 9

how is the number of plaques written?

Answer 9

plaque forming units per mL in the original phage stock solution

Question 10

what happens on the plate if the concentration of phage is too high?

Answer 10

the plaques will merge into one another making it difficult to determine how many are present

Question 11

what happens on the plate if the concentration of phage is too low?

Answer 11

it cannot be used to get an accurate estimate of the phage concentration in the stock solution

Question 12

how do we obtain an appropriate concentration of phage?

Answer 12

a series of dilutions of the phage stock solution are prepared, and then a known volume of each phage dilution solution (mixed with E coli B) is spread onto a series of tryptone agar plates

Question 13

plates with what range of phages are usually chosen for counting?

Answer 13

25-250

Question 14

describe the serial dilution procedure

Answer 14

5 test tubes each with 9mL saline. 1ml phage solution was added to the first one and mixed. 1ml of that solution was added to the next tube, and so on. Tubes were 10^-1, up to 10^-5

Question 15

describe the infection of bacteria and plating

Answer 15

added E coli to 5 microfuge tubes, and added phage solution. After incubation, each concentration of mixture was added to its own plate

soft tryptone agar was poured overtop and swirled to coat the dish