You are attempting to prepare a single gene knockout library using the pRL27 transposon system. You grow the donor E. coli in Luria broth containing both kanamycin and diaminopimelic acid (DAP) and your recipient Serratia rubidaea in plain Luria broth. You combine an equal ratio of donor and recipient cultures and plate the mixture onto Luria agar supplemented with DAP. After 24 hours incubation at 37°C, you create a cell slurry and plate the cells onto Luria agar aupplemented with kanamycin. After 24 hours incubation at 37°C, you find that no colonies grow. What best explains this outcome? A. Failure to supplement media with DAP B. Failure to remove antiobiotic containing media C. Failure to incubate for a sufficient length of time D. Failure to incubate at the appropiate temperature E. Failure to use the proper mating mix ratio

You are attempting to prepare a single gene knockout library using the pRL27 transposon system. You grow the donor E. coli in Luria broth containing both kanamycin and diaminopimelic acid (DAP) and your recipient Serratia rubidaea in plain Luria broth. You combine an equal ratio of donor and recipient cultures and plate the mixture onto Luria agar supplemented with DAP. After 24 hours incubation at 37°C, you create a cell slurry and plate the cells onto Luria agar aupplemented with kanamycin. After 24 hours incubation at 37°C, you find that no colonies grow. What best explains this outcome? A. Failure to supplement media with DAP B. Failure to remove antiobiotic containing media C. Failure to incubate for a sufficient length of time D. Failure to incubate at the appropiate temperature E. Failure to use the proper mating mix ratio

Human Heredity: Principles and Issues (MindTap Course List)

11th Edition

ISBN:9781305251052

Author:Michael Cummings

Publisher:Michael Cummings

Chapter13: An Introduction To Genetic Technology

Section: Chapter Questions

Problem 20QP: Analyzing Cloned Sequences A base change (A to T) is the mutational event that created the mutant...

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Genomic DNA from a family where sickle-cell disease is known to be hereditary, is digested with the restriction enzyme MstII and run in a Southern Blot. The blot is hybridised with two different 0.6 kb probes, both probes (indicated in red in the diagram below) are specific for the β-globin gene (indicated as grey arrow on the diagram below). The normal wild-type βA allele contains an MstII restriction site indicated with the asterisk (*) in the diagram below; in the mutated sickle-cell βS allele this restriction site has been lost. What size bands would you expect to see on the Southern blots using probe 1 and probe 2 for an individual with sickle cell disease (have 2 βS alleles)? Probe 1 Probe 2 (a) 0.6kb 0.6kb and 1.2kb (b) 0.6kb and 1.8kb 0.6kb, 1.2kb and 1.8kb (c) 1.2kb 0.6kb (d) 1.8kb 1.8kb a. (a) b. (b) c. (c) d. (d)
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T. Miyake and M. Demerec examined proline-requiring mutations in the bacterium Salmonella typhimurium (). On the basis of complementation testing, they found four proline auxotrophs: proA, proB, proC, and proD. To determine whether proA, proB, proC, and proD loci were located close together on the bacterial chromosome, they conducted a transduction experiment. Bacterial strains that were proC+ and had mutations at proA, proB, or proD were used as donors. The donors were infected with bacteriophages, and progeny phages were allowed to infect recipient bacteria with genotype proC− proA+ proB+ proD+. The recipient bacteria werethen plated on a selective medium that allowed only proC+ bacteria to grow. After this, the proC+ transductants were plated on selective media to reveal their genotypes at the other three pro loci. The following results were obtained: Q.Which genotypes represent single transductants and which represent cotransductants?
T. Miyake and M. Demerec examined proline-requiring mutations in the bacterium Salmonella typhimurium (). On the basis of complementation testing, they found four proline auxotrophs: proA, proB, proC, and proD. To determine whether proA, proB, proC, and proD loci were located close together on the bacterial chromosome, they conducted a transduction experiment. Bacterial strains that were proC+ and had mutations at proA, proB, or proD were used as donors. The donors were infected with bacteriophages, and progeny phages were allowed to infect recipient bacteria with genotype proC− proA+ proB+ proD+. The recipient bacteria werethen plated on a selective medium that allowed only proC+ bacteria to grow. After this, the proC+ transductants were plated on selective media to reveal their genotypes at the other three pro loci. The following results were obtained: Q.Is there evidence that proA, proB, and proD are located close to proC? Explain your answer.
T. Miyake and M. Demerec examined proline-requiring mutations in the bacterium Salmonella typhimurium (). On the basis of complementation testing, they found four proline auxotrophs: proA, proB, proC, and proD. To determine whether proA, proB, proC, and proD loci were located close together on the bacterial chromosome, they conducted a transduction experiment. Bacterial strains that were proC+ and had mutations at proA, proB, or proD were used as donors. The donors were infected with bacteriophages, and progeny phages were allowed to infect recipient bacteria with genotype proC− proA+ proB+ proD+. The recipient bacteria werethen plated on a selective medium that allowed only proC+ bacteria to grow. After this, the proC+ transductants were plated on selective media to reveal their genotypes at the other three pro loci. The following results were obtained: Q.Why are there no proC− genotypes among the transductants?
You are engineering a new vector that contains a screenable marker that can be used for blue/white screening of successful clones. For each site (1, 2, and 3) on the cloning vector below, describe why it would or would not be a good place for you to put the polylinker to facilitate blue/white screening. You can assume that the polylinker itself will not interfere with coding sequence in that region. In other words, the polylinker length will be a multiple of 3 nucleotides, will not contain a stop codon, and any amino acids translated will not affect the activity of the protein in that region. The arrows indicate the direction of transcription for the gene.
You are engineering a new vector that contains a screenable marker that can be used for blue/white screening of successful clones. For each site (1, 2, and 3) on the cloning vector below, describe why it would or would not be a good place for you to put the polylinker to facilitate blue/white screening. You can assume that the polylinker itself will not interfere with coding sequence in that region. In other words, the polylinker length will be a multiple of 3 nucleotides, will not contain a stop codon, and any amino acids translated will not affect the activity of the protein in that region. The arrows indicate the direction of transcription for the gene. Note from student:As stated in the problem... "YOU CAN ASSUME THAT THE POLYLINKER ITSELF WILL NOT INTERFERE WITH CODING SEQUENCE IN THAT REGION. IN OTHER WORDS, THE POLYLINKER LENGTH WILL BE A MULTIPLE OF 3 NUCLEOTIDES, WILL NOT CONTAIN A STOP CODON, AND ANY AMINO ACIDS TRANSLATED WILL NOT AFFECT THE ACTIVITY OF THE PROTEIN IN THAT…
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Analyzing Cloned Sequences A base change (A to T) is the mutational event that created the mutant sickle cell anemia allele of beta globin. This mutation destroys an MstII restriction site normally present in the beta globin gene. This difference between the normal allele and the mutant allele can be detected with Southern blotting. Using a labeled beta globin gene as a probe, what differences would you expect to see for a Southern blot of the normal beta globin gene and the mutant sickle cell gene?
rs1800092 is a single nucleotide variant identifier for a variant in the CFTR gene. What type of variant is it? What is the amino acids change? Is it pathogenic?