Genetics, Genomics, and Basic Definitions

• The material covers Genetics, Genomics, Chromosomes, Genotype, Phenotype, Semiconservative replication, Categories of Genes, Locations of genomes, DNA Replication, Transcription and Translation, Operons, Recombinant DNA, Horizontal Gene Transfer, Transposons, Mutations, Restriction enzymes, PCR, Gel Electrophoresis, Protein Structure, Antimicrobial Controls, Chemical and Physical methods of microbial control, and mechanisms and action of antimicrobial drugs.

Genetics, Genomics, and Basic Definitions

• Genetics is the study of inheritance and heredity in living organisms involving transmission from parent to offspring, trait expression, and structural/functional changes.
• Genomics involves studying an organism's entire genome, including interactions within the genome and the environment.
• The term "genome" refers to an organism's total genetic material, typically chromosomes, plasmids, as well as mitochondria and chloroplasts.
• Chromosomes are distinct structures composed of a neatly packaged DNA molecule.
• In eukaryotes, DNA is wrapped around histone proteins and is linear, existing in diploid or haploid states within the nucleus.
• In bacteria, DNA condenses into a packet using histone-like proteins, forming one, two, or sometimes several chromosomes.
• A "gene" represents the fundamental physical and functional unit of heredity, constructed from DNA, and containing instructions required for protein synthesis.
• Genotype refers to the sum of all gene types constituting an organism's unique genetic makeup.
• Phenotype is the expression of the genotype, yielding specific traits such as structures or functions.
• Semiconservative replication is a DNA replication method where each resulting DNA molecule consists of one original and one newly synthesized strand.

Categories and Locations of Genes

• Structural genes code for proteins.
• Regulatory genes control gene expression.
• There are genes that code for RNA used in protein production.
• Genomes are located in the nucleus within eukaryotes.
• In prokaryotes, genomes are located in the cytoplasm.
• In eukaryotic organelles, genomes are located in the mitochondria (matrix) and chloroplasts (stroma).
• Viruses contain DNA or RNA enclosed inside a protein coat known as a capsid.

DNA Structure, Replication, and Enzymes

• DNA strands run in a 5' to 3' direction and are antiparallel with one side of the helix oriented in the opposite direction compared to the other.
• One DNA strand runs from 5' to 3', while its complementary strand runs from 3' to 5'.
• DNA polymerase adds nucleotides to a growing DNA chain in the 5' to 3' direction.
• Nucleotides are joined together by covalent bonds to form the DNA backbone.
• Nitrogenous bases attach to the DNA strand at the 1' position of the deoxyribose sugar via a covalent bond.
• Purines and pyrimidines form complementary base pairs using hydrogen bonds.
• Purines are larger, two-ring structures designated with shorter names.
• Pyrimidines are smaller, one-ring structures designated with longer names.
• A purine must pair with a pyrimidine because two purines would be too wide, while two pyrimidines would be too narrow.
• Adenine (A) always pairs with Thymine (T).
• Guanine (G) always pairs with Cytosine (C).
• Helicase unzips the DNA helix.
• DNA Polymerase III adds new nucleotides to the DNA chain and proofreads for mistakes.
• Primase adds an RNA primer to initiate DNA synthesis.
• Ligase seals gaps and nicks in DNA during synthesis and repair.
• Gyrase helps untangle DNA supercoils.
• DNA Polymerase I removes the primer, fills gaps, and repairs mismatches.
• Topoisomerases I and II are involved in supercoiling and untangling DNA.

Transcription, Translation & Genetic Code

• Messenger RNA (mRNA) carries genetic information.
• Transfer RNA (tRNA) brings amino acids to ribosomes.
• Ribosomes are composed of small and large subunits and are the site of protein synthesis.
• A small ribosomal subunit binds to mRNA.
• A large ribosomal subunit forms peptide bonds between amino acids.
• Codons are groups of three nucleotides that specify which amino acid is added to the growing peptide chain.
• There are 64 different triplet codes.
• There are 20 amino acids.
• Some amino acids are represented by multiple codons, allowing for redundancy and correct amino acid insertion.
• Wobble refers to the fact that mainly the first two nucleotides of a codon are required to encode the correct amino acid.
• Anticodons are loops on tRNA that are complementary to mRNA codons.

Operons: Gene Regulation

• Operons are present in bacteria and archaea and regulate genes as a single unit.
• Operons can be inducible or repressible.
• Catabolic operons are induced by the substrate of the enzymes for which structural genes code.
• Repressible operons are anabolic enzymes, turned off by the product of the synthesis.

Horizontal Gene Transfer and Transposons

• Recombinant DNA (rDNA) is artificially made DNA formed via a combination of gene sequences.
• Conjugation is direct DNA transfer via a pilus from a donor to a recipient cell commonly transferring drug and metals resistance, toxin production, and enzyme adherence molecules.
• Transformation is the uptake of foreign DNA from the environment by a live, competent recipient cell, using polysaccharide capsules.
• Transduction involves bacteriophages transferring DNA from a donor to a recipient cell.
• Transposons are "Jumping Genes", transposable elements capable of shifting from one part of the genome to another.
• Transposons can be transferred from a chromosome to a plasmid, or vice versa.
• Transposons can be transferred from one cell to another in bacteria and some eukaryotes.
• Some transposons replicate before moving, while others simply move locations.
• Transposons can cause changes in traits, replace damaged DNA, and transfer drug resistance.
• A transposon may excise itself and move from one location to another in the genome, remaining at a single copy.
• A transposon may replicate before moving, increasing its copy number and impact on the host genome.
• A transposon may jump to a plasmid, which can then be transferred to another bacterial cell.

Mutations and Restriction Enzymes

• Spontaneous mutations are random changes in DNA arising from replication errors.
• Induced mutations result from exposure to mutagens like UV light, X-rays, and nitrous acid, disrupting DNA.
• Point mutations involve addition, deletion, or substitution of single bases, including missense, nonsense, silent and frameshift mutations.
• In point mutations, the wild type is the normal version of a gene.
• Missense mutations involve a change in the code leading to placement of a different amino acid, creating a faulty protein or no significant alteration.
• Nonsense mutations change a normal mutation into a stop codon.
• Silent mutations alter a base with no effect.
• Back-mutations occur when a mutated gene reverses to its original base composition.
• Frameshift mutations involve insertion or deletion of bases, altering the mRNA reading frame resulting in a nonfunctional protein.
• Restriction endonucleases recognize and cleave foreign DNA.
• Restriction endonucleases protect bacteria, and allow biotechnologists to cleave DNA at desired sites, which is important for recombinant DNA technology.
• Restriction endonucleases recognize and clip at palindromes.
• Palindromes are identical DNA sequences when read in the 5' to 3' direction on both strands.
• Restriction fragments are DNA pieces produced by restriction endonucleases.
• Restriction Fragment Length Polymorphisms (RFLPs) are differences in restriction endonuclease cutting patterns.

Other Enzymes, PCR, and Gel Electrophoresis

• Ligase seals sticky ends together and is used for splicing genes into plasmids.
• Reverse transcriptase (RT) converts RNA into DNA.
• Reverse transcriptase plays a key role in AIDS virus replication.
• Complementary DNA (cDNA) is made from messenger, transfer, ribosomal, and other forms of RNA.
• Complementary DNA synthesizes eukaryotic genes from mRNA transcripts.
• Polymerase Chain Reaction (PCR) rapidly increases the amount of DNA in a sample without culturing techniques.
• Polymerase Chain Reaction is sensitive enough to detect cancer from a single cell or diagnose an infection from a single gene copy.
• A PCR thermal cycler automatically performs cyclic temperature changes during PCR.
• Taq polymerase isolated from Thermus aquaticus remains active at elevated temperatures during PCR.
• Primers are DNA strands 15 to 30 bases long that serve as landmarks for DNA amplification.
• Denaturation occurs at 94 degrees C during PCR, to separate double-stranded DNA.
• Priming occurs at 50 -65 degrees C during PCR, allowing primers to bind to the target.
• Extension (Elongation) occurs at 72 degrees C during PCR , enlisting DNA polymerase to add nucleotides.
• The number of DNA doubles is repeated in cycles within 2, 4, 8, 16 steps.
• Gel electrophoresis separates DNA fragments with different lengths (large = slow, small = fast).
• Gel electrophoresis needs electrical current.
• Gel electrophoresis produces DNA fragments of different lengths in digested samples.
• Samples are placed in soft agar gel and subjected to an electrical current.
• Phosphate groups have a negative charge causing DNA to move toward the positive pole in the gel.
• Larger fragments migrate slowly while smaller fragments migrate quickly.
• The position of fragments is determined by staining the gel.

Protein Structure and Antimicrobial Controls

• Protein folding determines function.
• Protein shape and function correspond.
• Protein folding develops through primary, secondary, tertiary, and quaternary structures.
• Primary structure is the amino acid sequence which affects order and number of amino acids.
• Amino acids are linked together, forming one or more polypeptide chains.
• Secondary structure occurs with sequence of amino acids in the primary structure; alpha helix and beta pleated sheets.
• Tertiary structure occurs via a 3D shape when further folding occurs due to R groups (side chains).
• Hydrophilic R groups are external; hydrophobic R groups are internal.
• Quaternary structure is a protein comprised of more than one polypeptide chain.
• Sterilization eliminates viruses and viable microorganisms, and is used for inanimate objects.
• Disinfection is the destruction or removal of vegetative pathogens used on Inanimate objects, but not endospores- uses heat, bleach, iodine.
• Decontamination, or sanitization involves cleansing to lower contamination levels- uses detergents, soaps and dishwashers.
• Antisepsis, or degermation, destroys or inhibits vegetative pathogens on body surfaces; using scrubbing or submersion in chemicals like scrubbing with alcohol or hand sanitizer.

Relative Resistance and Germicides

• Microorganisms with the capacity to cause infection or spoilage in the environment are the primary targets of microbial control.
• Endospores are the most resistant microorganisms and is the goal to any sterilization treatment.
• Any procedure that eliminates endospores will eliminate all germ types.
• Agents denoted with "static" or "stasis" inhibit growth or stand still.
• Bacteriostatics, fungistatics, and antiseptics are microbistatic.
• Sterilization and disinfection is the process.
• Bactericides, fungicides, virucides, and sporicides are agents of the process.
• Sepsis is the growth of microorganisms in blood and tissues.
• Asepsis prevents microorganism entry into tissues.
• Permanent termination of vital processes defines microbial death.
• Thermal death time (TDT) is the shortest time to kill microbes at a certain temperature.
• Thermal death point (TDP) is the lowest temperature needed to kill all microbes for 10 minutes.
• A germicide is used for chemical controls, with rapid action, alcohol/water solubility, and affordability.
• Germicide application and control relies on the nature of the microbes being treated and of the product, as well as the exposure time, degree of contamination, and strength.
• Antimicrobial medications are described as germicides against certain bacteria strains and work by means of heat, radiation, or chemicals - these processes work by targeting the surface, cytoplasmic membrane, cell synthesis, or proteins.
• Spores and some bacteria are killed or affected by halogens, acids and alkalis, detergents, steriliants, and ethylene oxides.

Action of Chemotherapy

• Antimicrobial chemotherapy aims to administer a drug to an infected person to eliminate the agent without harming the host.
• Chemotherapy aims to disrupt an organism's structure or function so it can't survive; must be stable and easily moved and assimilated into the patient, while also being nontoxic.
• This method has 5 drug categories to target cell wall synthesis, structure, protein synthesis, cytoplasmic membrane structure function, or folic acids
• Prophylaxis, antimicrobial, antibiotic, semisynthetic, and synthetic drugs are antimicrobial.
• A narrow or broad spectrum for the drugs determine whether they help specifically or generally.
• Disc measuring is used to compare a strain and agent with the Kirby Bauer test.
• The therapeutic index is the ratio measuring the dose to the harm compared to what is required; small ratio is toxic, a higher one is better.
• Selective toxicity limits the cells toxicity.
• These treatments all target structure and function.
• All target either DNA, RNA, folic acids.
• A limited or wide variety affects the spectrum.
• Fungi are similar and harmful.
• Drug groups include polymixins, quinines, and antifungals like azole, metronidazoles.

Drug Resistance and Threats

• An adaptive response in which microorganisms begin to tolerate an amount of drug that would normally be inhibitory is referred to as drug resistance.
• New resistance can develop via spontaneous mutations.
• Slowing or stopping metabolism can also lead to microbial resistance as well as metabolic mutations
• Mechanisms behind drug resistance include producing enzymes that inactivate the drug and shutting down the cell.
• Due to such an increase, we may face a "post anabiotic era".
• The aim is that the drugs cause a threat to any or all of the organs.