DNA and RNA

• Nucleic acids are macromolecules essential for life, including DNA and RNA.
• Nucleotides, composed of a sugar, phosphate group, and nitrogenous base, are the monomers of nucleic acids.

DNA (Deoxyribonucleic Acid)

• The sugar in DNA is deoxyribose.
• The nitrogenous bases in DNA are adenine (A), thymine (T), cytosine (C), and guanine (G).
• The order of nucleotides determines specificity.
• DNA is a double-stranded molecule that forms a double helix structure, resembling a spiral staircase.
• The sides of the DNA staircase consist of deoxyribose and phosphate groups.
• Each step comprises two nitrogenous bases: A pairs with T, and C pairs with G.
• The two DNA strands are anti-parallel, with one strand having a terminal phosphate group (5' end) and the other a terminal hydroxyl group (3' end).
• DNA contains hereditary information and acts as the genetic blueprint.

RNA (Ribonucleic Acid)

• The sugar in RNA is ribose.
• The nitrogenous bases in RNA are adenine (A), uracil (U), cytosine (C), and guanine (G).
• In RNA, A pairs with U, and C pairs with G.
• RNA is a single-stranded molecule.
• Three types of RNA exist:
  • Messenger RNA (mRNA): Acts as an intermediary.
  • Ribosomal RNA (rRNA): Forms ribosomes along with proteins.
  • Transfer RNA (tRNA): Transfers amino acids.

Nucleic Acid Key Differences

• DNA contains deoxyribose, RNA contains ribose
• DNA contains the bases A, T, C, and G whilst RNA contains A, U, C, and G
• DNA is 2 stranded and RNA is 1 stranded

Genome and Genes

• Genome: The complete genetic material of a cell or virus.
• Gene: A specific nucleotide sequence coding for a protein or RNA.
• Escherichia coli has about 4,300 genes; humans have around 20,000.

DNA Replication

• DNA replication is the process of copying DNA.
• It occurs before binary fission in bacteria.
• It is semi-conservative: each new DNA molecule contains one original and one new strand.
• DNA replication in prokaryotes starts at a single origin (ori site) and proceeds bidirectionally.
• Helicase unwinds and unzips the DNA at the ori site, requiring ATP.
• Single-stranded binding proteins prevent the DNA from re-zipping.
• DNA polymerase replicates DNA by adding complementary nucleotides at a rate of 1,000 nucleotides per second in the 5' to 3' direction.
• The leading strand is replicated continuously, while the lagging strand is replicated discontinuously in Okazaki fragments.
• Okazaki fragments are joined together by DNA ligase.
• E. coli replicates its chromosome in about 40 minutes.
• DNA polymerase proofreads the new DNA strands during replication.

Flow of Information in a Cell

• The central dogma of biology describes the flow of information: DNA → mRNA → protein.
• Transcription: DNA is converted to mRNA.
• Translation: mRNA is converted to protein.
• Prokaryotes transcribe and translate genes based on need (e.g., lactose metabolism enzymes).

Transcription

• In prokaryotes, transcription happens in the cytoplasm, whereas in eukaryotes, it takes place in the nucleus.
• Only one DNA strand acts as a template, resulting in a single-stranded mRNA.
• Helicase unwinds and unzips the DNA; RNA polymerase moves along the template strand.
• RNA polymerase adds complementary nucleotides (50 nucleotides per second) to build the mRNA strand, where A pairs with U.
• Transcription starts at the gene's promoter and ends at the gene's termination point.
• mRNA is released, and the DNA re-zips and re-twists.
• Rifampin is an antibiotic that prevents transcription by blocking RNA polymerase.

Translation

• Translation occurs in the cytoplasm in both prokaryotes and eukaryotes.
• Ribosomes, the site of protein synthesis, are required.
• Prokaryotes have 70S ribosomes (30S and 50S subunits), while eukaryotes have 80S ribosomes (40S and 60S subunits).
• tRNA brings amino acids to the ribosome; amino acids are the building blocks of proteins.
• The ribosome moves along the mRNA, reading it three nucleotides (a codon) at a time.
• Codons specify an amino acid; anticodons on tRNA are complementary to mRNA codons, ensuring correct amino acid order.
• Streptomycin and tetracycline antibiotics prevent translation by binding to the 30S subunit of prokaryotic ribosomes.
• Degeneracy in the genetic code: multiple codons can encode the same amino acid.
• Mistakes in the 3rd position (wobble position) of the mRNA codon may not affect the resulting protein.

Genetic Engineering

• Genetic engineering involves altering an organism's genetic material to change its traits or produce a biological product.
• Recombinant DNA: DNA from two or more organisms spliced together, such as human and E. coli DNA.
• Restriction enzymes cut DNA at specific nucleotide sequences and are essential for producing recombinant DNA.
• Restriction enzymes can cut a human gene from human DNA.
• Plasmids from E. coli can be opened with the same restriction enzyme.
• The human gene can be inserted into the plasmid and sealed with DNA ligase.
• The plasmid, containing the human gene, is put inside of E. coli via transformation.
• E. coli will then transcribe and translate the human gene.
• Uses:
  • Production of human proteins like insulin, clotting factors, interferon, and growth hormone.
  • Increases milk production with recombinant bovine growth hormone.
  • Manipulates organisms for antibiotic production, herbicide/pesticide resistance, and vaccine production (e.g., hepatitis B and Gardasil).

DNA Fingerprints

• Restriction enzymes produce DNA fingerprints for crime scene analysis and paternity testing.
• Restriction fragment length polymorphism (RFLP): DNA is cut into a unique number of fragments due to individual differences in nitrogenous base order.
• Gel electrophoresis separates DNA fragments by size to create a pattern.
• The DNA pattern of a suspect must match crime scene samples for a guilty conviction.
• Children inherit half their DNA from each parent.

Polymerase Chain Reaction (PCR)

• PCR multiplies DNA, creating billions of exact copies from a single copy within hours.
• Applications:
  • Crime scene analysis (if the sample is too small).
  • Diagnostic testing (HIV, West Nile virus, Lyme disease)

Gene Therapy

• Gene therapy corrects defective genes responsible for disease development by inserting a normal gene to replace a disease-causing one.
• Vectors deliver therapeutic genes to target cells, with viruses being the most common vector.
• Viral vectors unload their genetic material, including the therapeutic human gene, into target cells.
• The gene can be introduced directly into the cell
• Liposomes can be used to carry the gene into the cell
• An artificial chromosome can be introduced into the cell along side the existing chromosomes
• Gene therapy faces numerous challenges:
  • Short-lived therapeutic DNA.
  • Immune system responses.
  • Viral vector toxicity and control issues.
  • Best suited for single-gene disorders.
  • Multi-gene disorders are difficult to treat effectively.

Human Genome Project (HGP)

• The HGP was an international research program aimed at completely mapping and understanding all human genes (the genome).
• Researchers determined the sequence of DNA bases, mapped gene locations on chromosomes, and produced linkage maps to track inherited traits.
• The HGP revealed approximately 20,500 human genes.
• The first draft of the human genome was published in February 2001, and the full sequence was completed in April 2003.
• The director of NHGRI noted that the genome is a history book, a shop manual, and a transformative textbook of medicine.