Module 20 - Plaque Assay of Phage
Call Kai
Learn
Practice Test
Spaced Repetition
Match
Flashcards
Knowt Play1/64
There's no tags or description
Looks like no tags are added yet.
Name | Mastery | Learn | Test | Matching | Spaced | Call with Kai |
|---|
No analytics yet
Send a link to your students to track their progress
65 Terms
What is a plaque assay used to determine?
The concentration of bacteriophage
What question does this lab answer?
How many viruses are in a sample
How do bacteriophages cause lysis of host bacteria?
Through the lytic cycle
What causes plaque formation in a bacterial lawn?
Lysis of bacteria by bacteriophages
What does PFU stand for?
Plaque-forming units
What does PFU measure?
The number of infectious virus particles
What does PFU/mL represent?
The concentration of phage in a sample
Why do plaques increase in size over time?
Because newly released phages infect neighboring cells, causing more lysis
What is a bacteriophage?
A virus that infects and replicates within bacteria
What phage is used in this lab?
T4 bacteriophage
What is another name for T4 bacteriophage?
Coliphage
Why is T4 called a coliphage?
Because it infects Escherichia coli
What type of bacterium is Escherichia coli?
Gram-negative bacterium
How is the phage introduced to bacteria in this lab?
By mixing phage suspension with a broth culture of bacteria
What type of agar is used in the overlay?
Soft or semisolid agar
What type of agar is used as the base?
Solid agar
What happens during the lytic cycle?
The virus takes over the host cell machinery to make new viruses
How does the virus reproduce inside the host cell?
By using host enzymes and ribosomes to make viral components
What happens after viral components are produced?
They assemble into complete virions
What happens to the host cell at the end of the lytic cycle?
It lyses (bursts), releasing new viruses.
What is a plaque?
A clear zone in a bacterial lawn where cells have been lysed
What causes a plaque to form?
Infection and lysis of bacteria by phages
What is a plaque-forming unit (PFU)?
A unit representing one infectious virus particle
What does one PFU originate from?
One virion infecting one bacterial cell
Why are plaques clear?
Because bacterial cells have been destroyed
What formula is used to calculate phage concentration?
PFU/mL = # of PFU/ (Volume of virus suspension plated x Dilution)
What must you know to calculate PFU/mL?
Number of plaques, volume plated, and dilution
What are viruses considered?
Obligate intracellular parasites
Why are viruses obligate intracellular parasites?
They must use host cell machinery to reproduce
What are viruses that infect humans called?
Animal viruses
What are viruses that infect bacteria called?
Bacteriophages
How are virus dilution tubes labeled?
V1, V2, V3
How are E. coli tubes labeled?
E1, E2, E3
How are plates labeled?
#1 (10^-2), #2 (10^-3), #3 (10^-4)
How much TSB is added to V1?
9.9 mL
How much TSB is added to V2?
0.9 mL
How much TSB is added to V3?
0.9 mL
How much E. coli culture is added to each E tube?
0.3 mL
What volume of phage is added to V1?
0.1 mL
What dilution is V1?
10^-2
How is V2 made?
Transfer 0.1 mL from V1 into V2
What dilution is V2?
10^-3
How is V3 made?
Transfer 0.1 mL from V2 into V3
What dilution is V3?
10^-4
How much virus is added to each E tube?
0.1 mL
Which tubes are mixed together?
V1 with E1, V2 with E2, V3 with E3
How long are tubes left for adsorption?
15 minutes
What happens during adsorption?
Phages attach to and penetrate host cells.
What is added to each E tube before plating?
Molten soft agar
What plate corresponds to E1?
#1 (10^-2)
What plate corresponds to E2?
#2 (10^-3)
What plate corresponds to E3?
#3 (10^-4)
What indicates successful phage replication?
Clear plaques in the bacterial lawn
What is a countable PFU range?
30â300 plaques
What does TNTC mean?
Too Numerous To Count
Why are counts below 30 not valid?
They are not statistically reliable
What is recorded for each plate?
Number of PFU, volume plated, dilution, and PFU/mL
What is the final goal of calculations?
Determine original phage density (PFU/mL)
Why is soft agar used instead of solid agar?
To allow phages and bacteria to spread and interact
What is a plaque?
A clear area caused by bacterial lysis
What is a PFU?
A measure of infectious virus particles
How are plaques and PFUs formed?
From infection, replication, and lysis of bacteria by phages
How do you record plates with fewer than 30 PFU?
As not statistically valid
How do you record plates with more than 300 PFU?
As TNTC
Why is a 15-minute adsorption period important?
To allow phages to attach to host bacteria before plating.