Laboratory Experiments in Microbiology (11th Edition)
11th Edition
ISBN: 9780321994936
Author: Ted R. Johnson, Christine L. Case
Publisher: PEARSON
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Chapter 32, Problem 1CA
Summary Introduction
To analyze:
Find the conclusion of given Ames test data.
Introduction:
Ames test is a screening process to identify high-risk compounds that are capable of causing mutations. It relies on using auxotroph of salmonella enterica, which cannot synthesize histidine (His-). The auxotroph has a mutation in dark excision repair (uvrB) and rfa.
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Let’s suppose you make a transposon library of the cellulose-secreting bacterium Komagataeibacter xylinus, with the goal of finding mutants that produce higher than normal amounts of cellulose, which would be useful industrially. However, despite your best efforts you are unable to isolate any transposon mutants that make more cellulose than the wild-type strain.Why might this have failed? List as many reasons as you can think of.
How would you modify the Ames test to evaluate physical mutagens?Would it be necessary to add the rat liver extract? Explain why or why not.
A classic way to isolate thymidylate synthase-negative mutants of
bacteria is to treat a growing culture with thymidine and trimetho-
prim. Most of the cells are killed, and the survivors are greatly
enriched in thymidylate synthase-negative mutants.
(a) What phenotype would allow you to identify these mutants?
(b) What is the biochemical rationale for the selection? (That is, why
are the mutants not killed under these conditions?)
(c) How would the procedure need to be modified to select mam-
malian cell mutants defective in thymidylate synthase?
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Laboratory Experiments in Microbiology (11th Edition)
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- What detection method could be used to determine the Mutation at a palindromic site and what results could be achieved from that?arrow_forwardA classic way to isolate thymidylate synthase-negative mutants of bacteria is to treat a growing culture with thymidine and trimethoprim. Most of the cells are killed, and the survivors are greatly enriched in thymidylate synthase-negative mutants. (a) What phenotype would allow you to identify these mutants? (b) What is the biochemical rationale for the selection? (That is, why are the mutants not killed under these conditions?) (c) How would the procedure need to be modified to select mammalian cell mutants defective in thymidylate synthase?arrow_forwardThe restriction enzymes KpnI and Acc65I recognize and cleave the same 6-bp sequence. However, the sticky end formed from KpnI cleavage cannot be ligated directly to the sticky end formed from Acc65I cleavage. Explain why.arrow_forward
- The restriction enzymes Kpn I and Acc 65I recognize and cleave the same 6-bp sequence. However, the sticky end formed from Kpn I cleavage cannot be ligated directly to the sticky end formed from Acc 65I cleavage. Explain why.arrow_forwardwhat is thermal stability and explain how the thermal stability of named proteins were improved using site directed mutagenesis.arrow_forwardAlthough a large number of mutagenic chemicals are known,none is known that induces mutations in only a single gene(gene-specific mutagenesis). From what you know aboutmutagens, explain why it is unlikely that a gene-specificchemical mutagen will be found. How then is site-specificmutagenesis accomplished?arrow_forward
- Silent mutations that occur in DNA are quite common in living cells and usually involve no effects onphenotype. In not more than 2 pages (using 1.5 line space of Arial or Times New Roman fonts) provideanswers for the following questions?1) Define the silent mutation in DNA? (2.5 marks)2) What is the codon usage bias? (2.5 marks)3) Provide one example of a clinical implication of a “silent mutation” that proven to have an effect onthe phenotype and provide a brief description of its molecular characteristics? (10.0 marks)arrow_forwardThe Ames test uses the reversion rate (His− to His+)to test compounds for mutagenicity.a. Is it possible that a known mutagen, like proflavin,would be unable to revert a particular His− mutantused in the Ames test? How do you think that theAmes test is designed to deal with this issue?b. Can you think of a way to use forward mutation(His+ to His−) to test a compound for mutagenicity? (Hint: Consider using the replica plating technique in Fig. 7.6.)c. Given that the rate of forward mutation is so muchhigher than the rate of reversion, why does the Amestest use the reversion rate to test for mutagenicity?arrow_forwardIn the Ames test shown in Figure 16-17, what is the reason for adding the liver extract to each sample?arrow_forward
- What is the purpose of Southern's blotting technique? Explain in detail the biochemical principle that underpin each step of the method.arrow_forwardExplain TWO (2) differences between two commonly used ligases; F. coli DNA ligase and T4 DNA ligase.arrow_forwardBased on the following wild type DNA sequence, indicate if each of the mutations should be classified as : insertion, deletion, missense, nonsense, silent (Use the provided Genetic Code table and remember you have been given DNA sequence). Wild Type: AUGAUUCUUAAAAGU Mutant 1: AUGAUUCUUUAAAGU Mutant 2: AUGAUUCUUGAAAGU Mutant 3: AUGAUCCUUAAAAGU Mutant 4: AUGAUCCUAAAAGU Mutant 5: AUGAUCCUUAAACAGU Socond letter Key: Ala = Alanine (A) Arg Arginine (R) Asn = UUU } UAU Tyr UGU UGC Cys UGA STOP UGG Trp UCU UCC UUC Phe Ser Asparagine (N) Asp = Aspartate (D) Cys Cysteine (C) Gin = Glutamine (Q) Glu = Glutamate (E) Gly = Glycine (G) His = Histidine (H) le = Isoleucine (1) Leucine (L) Lys Lysine (K) Met = Methionine (M) Phe = Phenylalanine (F) Pro Proline (P) Ser = Serine (S) Thr Threonine (T) Trp Tryptophan (W) Tyr Tyrosine (Y) - Valine (V) UCA UCG UAA STOP UAG STOP UUA Leu UUG S CCU CC CGU CUU CUC His CGC Arg Leu Pro CAA Gin CGA CCA CCG CUA CUG CGG Leu = AGU AUU AUC } lle AUA ACU ACC ACA Ser AAC…arrow_forward
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