Serial Dilutions & Plate Count Lab

Study Notes

Primary goal of serial dilutions and plate counts: Determine viable bacterial cell concentration in a sample.
Principle of plate counting: One bacterial cell forms one colony on an agar plate.
Creating a dilution: To create a 10⁻¹ dilution, transfer 1 mL of bacterial broth culture into 9 mL of sterile saline. To create a 10⁻² dilution, transfer 1 mL of the 10⁻¹ dilution into 9 mL of sterile saline.
Dilution factor calculation: Transferring 1 mL from a 10⁻⁶ dilution into 9 mL of sterile saline results in a final dilution factor of 10⁻⁷.
Calculating original bacterial concentration: Divide the number of colonies by the volume plated and multiply by the dilution factor. For example, 167 colonies from 0.1 mL of a 10⁻³ dilution gives ((167 colonies / 0.1 mL) * 10³) = 1.67 x 10⁶ viable cells/mL.

Spontaneous mutations: Random changes in DNA sequence that occur naturally during DNA replication. They can be beneficial, harmful or neutral for the microbe.
Media for assessing genetic diversity: YNB agar is used to assess genetic diversity in microbes.
Wild-type strain: A bacterial strain that exhibits the typical characteristics of its species.
Mutant strain: A bacterial strain that has a genetic alteration, resulting in a change in its phenotype.
Amino acid synthesis disruption: Mutant bacterial strains may lose the ability to synthesize a particular amino acid due to disruption in the biosynthetic pathway of that amino acid.

UV light damage on bacterial DNA: UV light forms pyrimidine dimers in DNA, which are covalent bonds between adjacent thymine or cytosine bases.
Effect of pyrimidine dimers on DNA base pairing: Pyrimidine dimers distort DNA structure and hinder proper base pairing during replication, leading to mutations or replication errors.
Lower UV killing rate in endospores vs. vegetative cells: Endospores have a lower rate of killing by UV light because they are less metabolically active and have DNA repair mechanisms.
Endospore resistance to UV light: The thick spore coat and lower metabolic activity of endospores contribute to their resistance to UV light.