BSCI223 - General Microbiology Exam II Review Notes

Bacterial growth can be characterized into four distinct phases: Lag phase, Log phase, Stationary phase, and Death phase.

Lag Phase
- Cells are not dividing but are metabolically active, preparing materials such as enzymes and machinery necessary for growth.
Log Phase (Exponential Growth)
- This phase is marked by rapid cell division as cells utilize available nutrients and multiply exponentially.
Stationary Phase
- Nutrient depletion occurs, and the rate of cell growth equals the rate of cell death, leading to a plateau in cell numbers.
Death Phase
- The rate of death surpasses the rate of growth, leading to a decline in the viable cell count and visible signs of cell lysis.

Turbidity Measurements
- Measure the cloudiness of the culture. Increased turbidity correlates with decreased transmittance and increased CFU/mL, calculated by measuring absorbance at OD(600).
Viable Plate Counts
- Conduct a series of 1:10 dilutions of the original culture, plating on agar to count colonies. Only plates with colony counts within the viable range of 30-300 should be used for calculations.
- Dilution Calculation:
  ext{Dilution} = rac{ ext{Initial Volume}}{ ext{Final Volume}}
  ext{Dilution Factor} = rac{ ext{Final Volume}}{ ext{Initial Volume}}
- For original culture concentration:
  ext{[Original Culture]} = rac{ ext{# colonies} imes ext{dilution factor}}{ ext{mL plated}}

Example Calculation:

If 65 colonies are counted from a 0.1 ml plated sample, then
ext{CFU/mL} = rac{65}{0.1} = 650 ext{ CFU/mL}
For a 10^{-3} dilution, the dilution factor is 1000, so:
$650 ext{ CFU/mL} imes 1000 = 650,000 ext{ CFU/mL} ext{ or } 6.5 imes 10^5$

Double Y Plot: Both CFU/mL and OD(600) values are displayed logarithmically.
Doubling Time: This is the time required for the bacterial population to double in number. It is analyzed and estimated from trendlines rather than raw data points.
Example time points indicate that for a doubling time of 32.5 min, a previous doubling event may have occurred at 25 min, resulting in 7.5 min as the time between them.

Defines the time for bacterial numbers to decrease by a factor of 10, gauged through survival percentage:
ext{% Survival} = rac{ ext{CFU/mL at time X}}{ ext{CFU/mL at time 0}}

Knowledge of correct plot reading and identifying inaccuracies is essential.
Growth curves involve calculating doubling time accurately using best-fit lines, and ensure that interactions between absorbance (OD) and CFU/mL are clearly understood.

Overview: The flow of genetic information proceeds from DNA to RNA to protein.

Transcription: Synthesis of RNA from DNA.
- Initiated at the promoter and terminated at the terminator. RNA Polymerase is the enzyme responsible for adding nucleotides in the 5' to 3' direction.
Translation: Synthesis of protein from RNA.
- Involves the Ribosomal Binding Site (RBS) and requires start and stop codons to signify the initiation and termination of protein synthesis.

DNA consists of nucleotides with adenine (A), thymine (T), guanine (G), and cytosine (C).
- Complementary Base Pairing: A pairs with T, linked by 2 hydrogen bonds, while C pairs with G with 3 hydrogen bonds.
- DNA is double-stranded and antiparallel (3' to 5' and 5' to 3').

Coding Strand (Sense Strand):
- Matches the RNA sequence, except T is replaced with U when transcribed.
Template Strand (Anti-Sense Strand):
- The strand that RNA Polymerase reads to create the mRNA transcript.

Mutations are heritable changes in the DNA sequence that can alter phenotypes or genotypes.
Sources can include DNA polymerase misincorporations, radiation exposure, and chemical mutagens.
Effects of mutations are categorized into:
1. Silent Mutations: Do not change the amino acid sequence (e.g., CAA to CAG).
2. Missense Mutations: Change one amino acid in the sequence (e.g., CAA to CAT).
3. Nonsense Mutations: Result in a premature stop codon (e.g., CAG to TAG) which often leads to a truncated protein.

Insertions or deletions (INDELs) that alter the reading frame if not in multiples of three can result in substantial changes to the encoded protein structure.

Used to evaluate mutagenic potential.
Detection of gain-of-function mutations in a bacterial strain (e.g., histidine-dependent Salmonella).
The experimental setup includes control and test groups, with significant growth observed in the test group indicating mutagenicity provided that the background mutation rates are low in the control.

An operon is a cluster of genes controlled by a single promoter, transcribed together into a single mRNA strand.
The lac operon is inducible and consists of genes necessary for lactose metabolism, including lacZ, lacY, and lacA.
The presence of lactose initiates transcription, while glucose can inhibit this process through catabolite repression, yielding a diauxic growth curve where glucose is used preferentially over lactose.

Transformation: Involves uptake of plasmid DNA by a competent bacterial cell.
Transduction: A bacteriophage injects DNA from one bacterium into another.
Conjugation: Direct transfer of DNA from a donor to recipient cell via pili, typically involving an F plasmid integrating into the host chromosome.

Characteristics of lytic and lysogenic cycles, with attention to the role of the prophage in dormancy versus lytic assembly and release.

Important components include origin of replication (ori), antibiotic resistance genes (blaA), screening genes (lacZ), and the regulatable promoter (lacI).
Different plasmids are constructed with varying efficiencies for purposes like gene expression in E. coli.