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Bio 139 Exam Review Outline: Exam #3 Ch. 7 DNA structure & function: Know functions of three RNA types (messenger, ribosomal, transfer). RNA polymerase: synthesizes RNA from a DNA template. What is a codon? How many nucleotides does it take to encode one amino acid?(3) What is the “genetic code” and how is it “degenerate”? Know that some codons mean “stop” (don’t need to memorize which ones). tRNAs have two functional ends: one binds an amino acid; the other has an anticodon. What does a tRNA do? What does a tRNA look like (shape)? Where does it work?(with a ribosome) What is a point mutation? How can it be “silent”? How can it produce a truncated (short) protein? (by introducing a stop codon) Insertion or deletion of nucleotide(s) not in a multiple of 3 causes frameshift. Why is a frameshift mutation usually more trouble than a point mutation? (changes entire subsequent protein sequence, not just one amino acid) Why is insertion/deletion of 3 or 6 nucleotides not as bad (as long as a stop codon isn’t introduced)? Briefly define enzymatic activities of nuclease/polymerase/ligase. Why is the Ames test used? What does it test for? (conversion of auxtroph to prototroph by random mutation, restoring the ability to synthesize an amino acid) What does it mean ?(a positive test, i.e., growth of colonies, indicates mutation has occurred and the test substance is mutagenic, and therefore is a potential carcinogen) Lac operon. Define constitutive vs inducible gene expression. Why is gene expression regulated at all? LacZ is a structural gene (what does that mean?) that codes for beta-galactosidase. What is a promoter? (regulatory gene where RNA polymerase binds and initiates transcription) Does its DNA sequence code for a protein? (no) What is an operator? (regulatory gene which controls expression of structural genes by serving as a sequence-specific binding site for DNA-binding proteins) Lac repressor gene (lacI) is always ON. Lac repressor protein binds to the lac operator and inhibits gene expression by blocking RNA polymerase. Repressor binds lactose (when present) and then loses the ability to bind to the operator, so expression (transcription & translation) of the lac genes turns ON. Be able to predict consequences of mutations in various components of the lac operon. Ask: can the cells utilize lactose? are the lac genes on or off? Is lacZ expression inducible or constitutive? PCR (polymerase chain reaction): repeated cycles of DNA synthesis to amplify a tiny amount of DNA. Done with temperature cycling to heat denature DNA & then synthesize new strands. Thermostable DNA polymerases are used (e.g., Taq) SKIP primers. Ch. 8 Recombinant DNA. Genetic code is universal. Restriction endonucleases cut DNA at specific sequences. They leave sticky ends (single strand overhangs). To express a foreign gene in bacteria you need a vector (like a plasmid) to carry it. To clone a gene, cut and paste the gene into a plasmid cut with the same restriction enzyme to create complementary sticky ends. Why is antibiotic selection important at the end? (the plasmid carries a resistance gene; bacteria carrying the plasmid are resistant; unwanted, nonrecombinant bacteria which do not have the plasmid with the cloned gene are killed). Ch. 13/14/15: antimicrobial spectrum of activity (broad vs narrow); problems with broad spectrum antibiotic use. Selective toxicity. Difference between drug toxicity & drug allergy. Normal microflora: What is it (dominant types, locations)? What is it good for? (keeping pathogens in check i.e. microbial antagonism; vitamin K synthesis) Define opportunistic infection. How can broad-spectrum antibiotics affect normal flora & cause disease? 2 classic superinfections (by the normal flora that follow antibiotic treatment of a different infection): C. dificile colitis; vaginal yeast (Candida). Antimicrobial mechanisms: 1. penicillins & peptidoglycan synthesis (requires beta lactam ring); 2. antifungal amphotericin B affects fungal membranes; 3. inhibiting 70S ribosome affects protein synthesis. 4. One example of an antimetabolite drug: sulfa drugs that competitively inhibit folic acid synthesis by mimicking PABA. Be able to state two mechanisms bacteria use to become antibiotic resistant. Steps toward establishing disease: access; attachment; colonization; invasion; damage Terms: virulence; attenuation. Exotoxin vs endotoxin. Intoxication vs infection. Incubation period. Zoonosis. Endemic, epidemic, pandemic. Emerging diseases. Common source vs propagated epidemic For bacterial diseases: pay attention to highlighted terms or facts; know names of organisms; pay attention to WHY I chose to present that particular pathogen (shows some special feature or aspect of pathogenesis, disease transmission, mechanism, etc.) #1 reason infectious disease deaths have declined dramatically in last 150 years: SANITATION. What are reservoirs of infection, and how are they important? Ch. 10. Viral structure: envelope (lipid bilayer), capsid (protein), nucleic acid core. Viral genomes: not always dsDNA like cells (all combinations of ss/ds RNA/DNA). How do enveloped viruses acquire a lipid bilayer membrane? What is an envelope good for, even though it makes the virus fragile? How do animal viruses enter a cell?(adsorption, then endocytosis, or membrane fusion for enveloped viruses). Unlike bacteriophage viruses which inject only DNA, animal viruses must be uncoated to expose the genome. RNA viruses: must provide enzymes for syntheses a cell can’t do. Understand difference between + and – sense RNA; how do they replicate? Any ssRNA virus must provide an RNA-dependent RNA polymerase enzyme (cells can’t do this). + sense RNA virus: brings the gene to manufacture such an enzyme; - sense RNA virus: must actually bring the enzyme with it inside the capsid. Retroviruses: totally different RNA viruses, make DNA (reverse transcription) & integrate as provirus. Antigenic drift vs Antigenic shift in influenza A viruses. Hemagglutinin & Neuraminidase: important surface antigens on the surface (envelope) of influenza that distinguish one strain from another