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Biol 240 Exam #2 Review Notes

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University of Waterloo
BIOL 240
Josh Neufeld

Topic 9 Textbook (pg. 285-) Mutation and Mutants Mutation = heritable change in the base sequence of that genome o Mutation brings a very small amount of genetic change in a cell o Genetic recombination typically generates much larger changes Mutation and genetic recombination fuel the evolutionary process Prokaryotes do not reproduce sexually (use other mechanisms for genetic exchange) In all cells (EXCEPT viruses), genome consists of double-stranded DNA o Virus can be single or double-stranded DNA or RNA Mutant = a strain of any cell/virus carrying a change in nucleotide sequence o Differs from parent in genotype o Can also have a mutant phenotype (observable properties) Genotype vs. Phenotype Genotype = designated by 3 lowercase letters followed by a capital letter o Ex. hisC gene of E. coli Phenotype = designed by a capital letter followed by 2 lowercase letters o Has a + or superscript to indicate the presence or absence of that property + - o Ex. His strain = can make histidine, His strain = cannot make histidine Isolation of Mutants: Screening vs. Selection Selectable mutation gives a clear advantage on the mutant strain under certain environmental conditions, so the progeny of the mutant cell are able to outgrow and replace the parent o Ex. drug resistance an antiobiotic-resistant mutant can grow in the presence of antibiotic concentrations that inhibit or kill the parent o Easy to detect and isolate selectable mutants (choosing diff. environmental conditions) Example of nonselectable mutation = colour loss in organism o Nonpigmented cells have no advantage/disadvantage over pigmented parent cells Screening = procedure that permits the identification of organisms by phenotype / genotype o Doesnt inhibit/enhance the growth of particular phenotypes or genotypes o detect mutations by examining large numbers of colonies (looking for different ones) Isolation of Nutritional Auxotrophs and Penicillin Selection 1 screening is more tedious than selection nutritionally defective mutants can be detected by the technique of replica plating auxotroph = mutant with a nutritional requirement for growth (a screening process) o prototroph = parent of auxotroph penicillin-selection method = used to isolate auxotrophs - o enrich a population of mutagenized cells in His mutants, then replica plating can be much more effecting (without enrichment, mutants will be rare in the culture) o penicillin is an antibiotic that kills only growing cells o if penicillin is added to a population of cells growing in a medium lacking the nutrient required by the desired mutant, the mutants will not grow and will therefore be unaffected by the penicillin; however, the parents will grow, and will therefore be killed o the resulting colonies will include some wild-type (that escaped killing by penicillin) and some desired mutants; therefore, penicillin doesnt necessarily select for the mutants, but selects against the parental type SUMMARY: penicilllin is often used as a prelude to replica plating to increase the chances of obtaining auxotrophic mutants (since mutants are normally rare in a mutagenized culture) 11.5 Molecular Basis of Mutation Induced mutation = mutations that are made deliberately Spontaneous mutation = mutations that occur without human intervention o can result from: exposure to natural radiation oxygen radicals errors in the pairing of bases during DNA replication (MAIN CAUSE) point mutations = mutations that change only 1 base pair o base-pair substitution or loss/gain of a single base pair o in coding region of gene that encodes a polypeptide Base-Pair Substitutions genetic code is degenerate = not all mutations in the base sequence encoding a polypeptide will change the polypeptide (ex. ABC could encode the same polypeptide as BDC) silent mutation = mutation that doesnt affect the phenotype of the cell o almost always in the 3 base of the codon (ex. TAC to TAT) 2 missense mutation = mutation in which a single codon is altered so that one amino acid in a protein is replaced with a different amino acid o ex. amino acid change from tyrosine to asparagines by changing UAC to AAC o may cause protein to be inactive or have reduced activity mutations in the active site of proteins are more likely to destroy activity nonsense mutation = mutation that changes a codon for an amino acid (sense) to a nonsense codon (causing a stop) o this may result in an incomplete product and will cause it to be inactive types of base substitution: transition and transversion o transition = mutation in which one purine base (A or G) is substituted for another purine, or one pyrimidine base (C or T) is substituted for another pyrimidine o transversion = point mutation in which a purine base is substituted for a pyrimidine base or vice versa purine = A or G pyrimidine = C or T Frameshifts and Other Insertions or Deletions since genetic code is read in blocks of 3 bases, any deletion/insertion of a single base pair results in a shift in the reading frame o can result from replication errors insertion/deletion of 1 or 2 base pairs causes a frameshift o messes up the entire polypeptide sequence after the mutation (read the wrong way) insertion/deletion of 3 base pairs either adds or removes 1 codon (which is = to 3 bases pairs) large insertions/deletions (of hundreds/thousands of base pairs) can result in complete loss of gene function and possible death o may be a result of errors during genetic recombination o many insertion mutations are due to insertion sequences (700-1400 bp in length) point mutations can be corrected through further mutation, large insertions/deletions cannot translocation = large section of DNA is moved to a new location inversion = orientation of a DNA segment is reversed relative to the surrounding DNA 3 Back Mutations or Reversions reversion = alteration of DNA that reverses effects of a previous (point) mutation o original phenotype that was changed in the mutant is restored o 2 types: Same-site revertant = mutation that restores activity at the same site as the original mutation; this is called a true revertant Second-site revertant = mutation that restores activity at a different site Can function as suppressor mutations o Suppressor mutation = mutation that compensates for the effect of the original mutation and restores the original phenotype; there are different classes 1. Mutation somewhere else in the same gene that restores enzyme function 2. Mutation in another gene that restores the function of the original mutant gene 3. Mutation in another gene that results in the production of an enzyme that can replace the mutated one Mutagenesis Mutagens = agent that causes mutation Chemical Mutagens Different classes of chemical mutagens: o Nucleotide base analogs = molecules that resemble the purine and pyrimidine bases of DNA in structure, yet display faulty pairing properties (ex. AT to GC substitutions) Causes DNA to replicate normally most of the time, but DNA replication errors still occur at higher frequencies at these sites due to incorrect pairing Incorrect pairing results in point mutations o Chemical modifications = results in faulty base pairing or related changes Ex. alkylating agents = able to introduce changes even in nonreplicating DNA o Intercalating agents = these mutagens are inserted between 2 DNA base pairs and in the process push them apart Can lead to insertions or delections in acridine-containing DNA Acradine typically induce frameshift mutations 4
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