Microbiology Lab (2065) Final Review. Fall 2023. SLCC

Virus properties (4)

1. They lack ribosomes (cannot metabolize on their own)

2. Have either DNA or RNA genome (not both)

3. Termed bacteriophage when infecting bacteria

4. Are smaller than bacteria

5. Surrounded by protein coat (capsid)

Virus definition

An obligate intracellular parasite that infect all cells: bacteria, animals, fungi, or plants

Virion

a subcellular infectious particle

Viral structure (basic parts)

• Nucleic acid

• Capsid (protein coat made of promoters or capsomeres)

• Spikes (carbohydrate-protein)

• Envelope (some)

• Naked viruses lack an envelope

viral structures

Helical viruses

Polyhedral (icosahedral) viruses

Enveloped viruses

complex

Prolate

What type of replication is either lytic or lysogenic?

Viral replication

Lytic replication

• Virus destroys host cell; includes virulent bacteriophages.

• Attachment, penetration, replication, assembly, release

Lysogenic replication

• Viral genome replicated along with host DNA due to integration

• Attachment, penetration, genome integration, environmental stimuli needed for lytic initiation (Virus remains in a "dormant" state until activated by an environmental trigger)

Temperate bacteriophages

• phages capable of using either "lytic" or "lysogenic" cycle

• Do not always destroy their host

On a micropipette, what does the red number indicate?

The decimal point/position relative to the pipette's total volume

In the plaque assay lab, what was used to infect the E. coli?

T4 phage

What equipment was used in the plaque assay lab?

PPE, Micropipette, incubator, vortexer, etc)

What is the control lane in gel electrophoresis?

Lane 1

What charge does DNA have?

Negative

In the gel electrophoresis, which pole will the charged DNA molecules move towards?

Positive pole

Which move slower in the gel matrix, longer or shorter DNA fragments?

longer fragments because they bump into the gel matrix more often

What plating technique is used with the plaque assay?

Pour

PFU

Plaque-Forming Units per milliliter. It is the concentration of bacteriophage stock

What number of plaques per plate is considered a valid phage count?

between 30 and 300

How is PFU calculated?

Average # of plaques / (dilution factor [we did 1] * (volume of diluted virus added [e.g. 1/10, or 1/100, or 1/1000])

in DNA sequencing, results given in FASTA format, what do all the "N"s mean if present?

At the particular position, the nucleotide was too degraded to call

E-values near zero mean what on the BLAST site?

results that were successful matches

on chromatograms, what direction does the nucleotide sequence read from?

5' to 3'

What are serial dilutions?

A dilution that is performed in progressive regular increments increased by a factor of 10. Each dilution is less concentrated than the other.

What is BLAST

Basic Local Alignment Search Tool; Finds regions of local similarity betwewen sequences.

Which type of plate would you expect to see the most growth concerning the complement experiment?

C plates

Why are PCR samples processed in gel electrophoresis before sending them off for sequencing?

• Ensure DNA was contained in PCR tubes

• Determine DNA size is contained in PCR tubes

• Separate DNA fragments based on size (shape) & charge

• determine if band indicating a good sample to send for sequencing.

Opsonization

Phagocyte recognition

Agglutination

Gathering of tagged molecules for destruction

Chemotaxis

Deployment of leukocytes

Cytolysis

Through MAC ring structure that creates pore in membrane

Does complement work on gram positive bacteria?

No; due to their thick cell wall.

What serum naturally contained complement that we combined with E. coli?

Bovine

What is the component added to the PCR reaction

Taq DNA polymerase

What life cycle does plaques happen at?

Death/decline phase