Microbiology Quiz Review: Bacterial Transformation, Ultraviolet Light, and Kirby-Bauer Method

Comprehensive review flashcards covering bacterial transformation (pGLO plasmid), ultraviolet light DNA damage and repair mechanisms, and the Kirby-Bauer disk diffusion assay protocol and interpretation.

What is bacterial transformation?

The process by which a competent bacterial cell takes up naked extracellular DNA from its environment and incorporates it into its own genome.

What is the function of the GFP (Green Fluorescent Protein) gene in the pGLO plasmid?

It acts as a reporter gene that produces green fluorescence under UV/blue light when expressed, visually confirming successful transformation.

What is the role of the bla gene in the pGLO plasmid?

It encodes beta-lactamase, providing resistance to ampicillin so only transformed bacteria survive on selective media.

How are bacterial cells made chemically competent to take up plasmid DNA?

Via calcium chloride and heat shock.

Which specific wavelength range of non-ionizing ultraviolet (UV) light causes microbial DNA damage?

UVC, specifically $200-290\,\text{nm}$.

How does ultraviolet (UV) light damage microbial DNA?

By causing adjacent thymine or cytosine bases to covalently link into pyrimidine dimers.

What structural impact do pyrimidine dimers have on microbial DNA?

They distort the DNA double helix, blocking normal DNA replication and transcription.

How does photoreactivation (light repair) repair UV damage in DNA?

Photolyase enzymes use visible light energy to split the pyrimidine dimers apart.

What is the mechanism of nucleotide excision repair (dark repair)?

Endonucleases excise the damaged DNA strand, and DNA polymerase and ligase resynthesize and seal the gap.

Why are bacterial endospores (such as Bacillus species) significantly more resistant to UV radiation than vegetative cells?

Due to protective small acid-soluble spore proteins (SASPs) and a dehydrated core.

What is the primary purpose of the Kirby-Bauer method?

It is a standardized disk diffusion assay used to determine the susceptibility or resistance of a bacterial isolate to various antimicrobial agents.

What growth medium and agar depth are required by the standardized Kirby-Bauer protocol?

Mueller-Hinton agar poured to a uniform depth of roughly $4\,\text{mm}$.

To what turbidity standard must the bacterial suspension be matched in the Kirby-Bauer method?

A $0.5$ McFarland turbidity standard.

What is the standard incubation time for the Kirby-Bauer disk diffusion test?

Overnight, typically $16-24\,\text{hours}$.

What are the clear areas around antibiotic disks in the Kirby-Bauer test called, and how are they measured?

Zones of inhibition, measured by their diameter in millimeters ($\text{mm}$).

What three classifications can an organism receive when comparing measured zones of inhibition to standard reference charts?

Resistant (R), Intermediate (I), or Susceptible (S).