Nucleic Acids: DNA and RNA Structures

• Nucleic acids include DNA, RNA, and polypeptides.

Nucleic Acid Structure

• Nucleotides consist of a phosphate group, a five-carbon sugar (deoxyribose in DNA, ribose in RNA), and a nitrogenous base.
• DNA contains deoxyribose sugar, while RNA contains ribose sugar.
• DNA nitrogenous bases: Adenine (A), Thymine (T), Cytosine (C), Guanine (G).
• RNA nitrogenous bases: Adenine (A), Uracil (U), Cytosine (C), Guanine (G).
• DNA is double-stranded (dsDNA); RNA is single-stranded (ssRNA).

Genetic Coding & Central Dogma

• Central dogma: DNA → RNA → Protein.
• DNA transcription yields RNA, which is translated into amino acids to form proteins.
• Base pairing rules: A-T in DNA, A-U in RNA, G-C in both DNA and RNA.

Pyrimidines vs. Purines

• Pyrimidines: Cytosine (C), Thymine (T) in DNA, Uracil (U) in RNA, and have one ring.
• Purines: Adenine (A), Guanine (G), and have two rings making them larger than pyrimidines.

DNA vs. RNA

• Similarities: Both are nucleic acids, possess sugar-phosphate backbones, and include A, C, and G bases.
• Differences: DNA is double-stranded, and RNA is single-stranded.
• DNA has deoxyribose, while RNA has ribose.
• DNA contains thymine (T), and RNA contains uracil (U).
• DNA is more stable, and RNA is more transient and participates in protein synthesis.

Types of RNA

• mRNA (Messenger RNA) carries genetic information from DNA to ribosomes.
• rRNA (Ribosomal RNA) forms the core of ribosomes and catalyzes protein synthesis.
• tRNA (Transfer RNA) brings amino acids to the ribosome during translation.

Transcription and Translation

• Transcription: DNA sequence is transcribed into a complementary RNA sequence using RNA polymerase.
• Translation: RNA sequence is translated into an amino acid sequence to create a polypeptide/protein.
• Codons, which are triplets of nucleotides, specify amino acids.

Template vs. Coding Strand

• Template strand: Used by RNA polymerase to synthesize mRNA.
• Coding strand: Identical to mRNA, except with T replaced by U in RNA.

IDEXX MPN System

• Colilert detects coliforms and E. coli; Enterolert detects Enterococcus.
• Color change or fluorescence indicates the presence of bacteria.
• Indicator organisms such as E. coli and Enterococcus signify fecal contamination.
• Antibiotics target bacterial structures, including the cell wall, ribosomes, and DNA replication enzymes.

Selective & Differential Media

• EC-MUG detects E. coli via fluorescence.
• EMB (Eosin Methylene Blue) differentiates lactose fermenters (E. coli exhibits a green metallic sheen).
• BE (Bile Esculin) identifies Enterococcus by hydrolyzing esculin.

Serial Dilutions, Heat-Shock, Antibiotic-Producing Bacteria, Plating

• Serial dilutions reduce bacterial concentration stepwise for quantification.
• Heat-shock involves exposing bacteria to high temperatures to select for heat-resistant strains.
• Antibiotic-producing bacteria naturally produce antibiotics (e.g., Streptomyces).
• Plating techniques for bacterial isolation and counting: streak plate, spread plate, pour plate.

Gram Stain Procedure

• Steps: Crystal violet (primary stain) → Iodine (mordant) → Alcohol wash (decolorization) → Safranin (counterstain).
• Results: Gram-positive bacteria stain purple due to a thick peptidoglycan layer retaining crystal violet.
• Gram-negative bacteria stain pink because of a thin peptidoglycan layer, losing crystal violet, and taking up safranin.

Quad-Streak & Isolation

• Quad-streak isolates pure bacterial colonies from a mixed culture by reducing bacterial density across four quadrants on an agar plate.

Media Used for Bacterial Culture

• Selective media suppresses unwanted microbes and selects for desired ones (e.g., EMB, MacConkey agar).
• Differential media distinguishes bacteria based on metabolic properties (e.g., EMB, blood agar).
• Enrichment media encourages the growth of specific microbes (e.g., nutrient broth for fastidious organisms).
• Maintenance media supports long-term bacterial storage (e.g., tryptic soy agar).