Question 1

How were GadE and GadX demonstrated to be transcription factors for the gadA gene?

Accepted Answer

Footprint analysis can be used to find out what nucleotides in a DNA fragment directly interact with DNA-binding proteins. The principle is that the DNA region bound by protein is protected from endonuclease activity (e.g., DNase I). After disruption of the protein-DNA interactions, protected DNA fragments are electrophoretically separated in a gel along with protein-free DNA that had been treated with DNase I under the same conditions. A set of sequencing reactions help in localizing the transcriptional start site and nucleotides that directly bind the protein, as only one of the DNA strands is labeled and migrates to a position that matches one base of the sequencing reactions. Both GadE and GadX protect several regions upstream of the transcriptional start of gadA.

Question 2

What is synthetic biology? Provide an example of its potential applications.

Accepted Answer

Synthetic biology is the conception and building of biological systems that do not exist in nature. This is accomplished by creating chimeric gene circuits that result in novel gene expression patterns and biochemical pathways. At the present time, genetic switches offer realistic capabilities of bacterial-based computing and the potential to use these circuits in the detection of toxins, explosives, and other nocive chemicals.

Question 3

His₆-fusion-tagged proteins can be recovered from affinity chromatography columns because the histidine tag tightly binds to beads coated with ________ ions. The elution buffer contains ________, which has stronger affinity for these ions.

Accepted Answer

A)  copper; sulfate 
B)  magnesium; EDTA 
C)  iron; citrate 
D)  calcium; EGTA 
E)  nickel; imidazole 
A)  copper; sulfate 
B)  magnesium; EDTA 
C)  iron; citrate 
D)  calcium; EGTA 
E)  nickel; imidazole E

Question 4

Noise in synthetic biology is a result of the

Accepted Answer

A)  ara operon. 
B)  lac operon. 
C)  dual-feedback oscillator. 
D)  fluctuation of gene expression among single-cells in a population. 
E)  oscillatory genetic circuit. 
A)  ara operon. 
B)  lac operon. 
C)  dual-feedback oscillator. 
D)  fluctuation of gene expression among single-cells in a population. 
E)  oscillatory genetic circuit.

Question 5

For a reporter protein to be synthesized, its gene must be fused in the proper codon reading frame with respect to the

Accepted Answer

A)  ribosome binding site. 
B)  promoter of the gene of interest. 
C)  vector origin of replication. 
D)  target gene. 
E)  vector's antibiotic marker sequence. 
A)  ribosome binding site. 
B)  promoter of the gene of interest. 
C)  vector origin of replication. 
D)  target gene. 
E)  vector's antibiotic marker sequence.

Question 6

Describe the difference between autoradiography and phosphor imaging.

Accepted Answer

In autoradiography, emissions from a rad

Question 7

Potential applications of bacterial oscillator switches include all of the following EXCEPT the

Accepted Answer

A)  study of controlled interactions between two feedback genetic loops. 
B)  design of bacterial genetic circuits to detect toxins. 
C)  design of bacterial clocks. 
D)  reduction of noise in biological circuits. 
E)  reactivity to concentrations of specific molecules such as arabinose. 
A)  study of controlled interactions between two feedback genetic loops. 
B)  design of bacterial genetic circuits to detect toxins. 
C)  design of bacterial clocks. 
D)  reduction of noise in biological circuits. 
E)  reactivity to concentrations of specific molecules such as arabinose.

Question 8

How do operon fusions reveal transcriptional control of a target gene?

Accepted Answer

Operon (transcriptional) fusions have a

Question 9

Which technique is used to select novel enzymes or peptides produced through directed evolution?

Accepted Answer

A)  mutagenesis with transposons 
B)  enzyme phage display 
C)  multiplex PCR 
D)  ethane methyl sulfonate mutagenesis 
E)  directional cloning 
A)  mutagenesis with transposons 
B)  enzyme phage display 
C)  multiplex PCR 
D)  ethane methyl sulfonate mutagenesis 
E)  directional cloning

Question 10

Which kind of phage vectors are most frequently used in phage display technology?

Accepted Answer

A)  double-stranded DNA T-odd 
B)  double-stranded DNA T-even 
C)  filamentous single-stranded DNA phages 
D)  lambda-based vectors 
E)  alpha-based vectors 
A)  double-stranded DNA T-odd 
B)  double-stranded DNA T-even 
C)  filamentous single-stranded DNA phages 
D)  lambda-based vectors 
E)  alpha-based vectors

Question 11

Sigma B is a stress-induced transcription factor in Bacillus subtilis. Bacteria producing a sigma B-YFP fusion protein were exposed to constant energy stress, and fluorescence pulses emitted by individual cells were detected by time-lapse microscopy. Could this result have been obtained using a sigma B-b-gal fusion protein?

Accepted Answer

A)  Yes. Hydrolysis of X-Gal by b-galactosidase produces a blue product detectable by eye. 
B)  Yes. Hydrolysis of ONPG by b-galactosidase in liquid medium produces an orange compound measurable by spectrophotometry. 
C)  Yes. It depends on how well the reporter gene is inserted into the vector with respect to the position of the ribosome-binding site. 
D)  No. Whereas YFP fluorescence can be carefully observed and measured by time-lapse microscopy, b-gal activity is measured as an average of the population as a whole. 
E)  No; the product would not be detectable by eye. 
A)  Yes. Hydrolysis of X-Gal by b-galactosidase produces a blue product detectable by eye. 
B)  Yes. Hydrolysis of ONPG by b-galactosidase in liquid medium produces an orange compound measurable by spectrophotometry. 
C)  Yes. It depends on how well the reporter gene is inserted into the vector with respect to the position of the ribosome-binding site. 
D)  No. Whereas YFP fluorescence can be carefully observed and measured by time-lapse microscopy, b-gal activity is measured as an average of the population as a whole. 
E)  No; the product would not be detectable by eye.

Question 12

The colorless molecule o-nitrophenyl galactoside (ONPG) can be used as an artificial substrate to assay b-galactosidase in cell extracts of Escherichia coli in which lacZ is used as a reporter gene. Which of the following statements is NOT true with respect to the use of ONPG?

Accepted Answer

A)  Hydrolysis of ONPG by b-galactosidase produces yellow o-nitrophenol. 
B)  o-nitrophenol can be measured with a spectrophotometer. 
C)  Hydrolysis of ONPG by b-galactosidase also releases colorless galactose. 
D)  The o-nitrophenol moiety is blue and more suitable for use in solid media. 
E)  The o-nitrophenol moiety is green and less suitable for use in solid media. 
A)  Hydrolysis of ONPG by b-galactosidase produces yellow o-nitrophenol. 
B)  o-nitrophenol can be measured with a spectrophotometer. 
C)  Hydrolysis of ONPG by b-galactosidase also releases colorless galactose. 
D)  The o-nitrophenol moiety is blue and more suitable for use in solid media. 
E)  The o-nitrophenol moiety is green and less suitable for use in solid media.

Question 13

Potential vaccine antigens made by transgenic plants include

Accepted Answer

A)  foot-and-mouth surface antigen. 
B)  Norwalk virus glycoprotein. 
C)  rabies virus glycoprotein. 
D)  Entertoxin A subunit. 
E)  Hepatitis A virus. 
A)  foot-and-mouth surface antigen. 
B)  Norwalk virus glycoprotein. 
C)  rabies virus glycoprotein. 
D)  Entertoxin A subunit. 
E)  Hepatitis A virus.

Question 14

The figure below depicts a "kill switch" or suicide module designed to eliminate an organism at a predetermined point, namely once the microorganism's function is accomplished. Explain how this kill switch can be regulated to prevent the untimely death of a microorganism harboring it.

Accepted Answer

CcdB is a powerful DNA damaging toxin. T

Question 15

How can a protein be detected on a western blot?

Accepted Answer

A sample of the protein of interest is i

Question 16

Use the figure below to describe how the CRISPR-Cas9 editing tool is used in eukaryotic cells.

Accepted Answer

[1] The Streptococcus pyogenes Cas9 endo

Question 17

Scientists have taken the genome of the bacterial species ________ and moved it to yeast.

Accepted Answer

A)  Mycobacterium leprosae 
B)  Mycobacterium tuberculosis 
C)  Mycoplasma mycoides 
D)  Escherichia coli 
E)  Mycoplasma pneuomoniae 
A)  Mycobacterium leprosae 
B)  Mycobacterium tuberculosis 
C)  Mycoplasma mycoides 
D)  Escherichia coli 
E)  Mycoplasma pneuomoniae

Question 18

Which probing technique detects DNA immobilized on a membrane using labeled DNA fragments?

Accepted Answer

A)  northern blot 
B)  southern blot 
C)  western blot 
D)  polyacrylamide gel electrophoresis in SDS 
E)  isoelectrofocusing 
A)  northern blot 
B)  southern blot 
C)  western blot 
D)  polyacrylamide gel electrophoresis in SDS 
E)  isoelectrofocusing

Question 19

CRISPR-Cas9, a prokaryotic defense mechanism, is a powerful emerging tool in biotechnology for all of the following EXCEPT

Accepted Answer

A)  allowing the editing of defective genes in eukaryotic cells. 
B)  introducing insertions, deletions, or sequence modifications into a given genome with CRISPR-Cas9. 
C)  regulating gene expression in bacteria with CRISPRi, in which case Cas9 is no longer active as a nuclease. 
D)  marking a defective gene with antibiotic resistance. 
E)  disrupting latent HIV provirus in infected cells. 
A)  allowing the editing of defective genes in eukaryotic cells. 
B)  introducing insertions, deletions, or sequence modifications into a given genome with CRISPR-Cas9. 
C)  regulating gene expression in bacteria with CRISPRi, in which case Cas9 is no longer active as a nuclease. 
D)  marking a defective gene with antibiotic resistance. 
E)  disrupting latent HIV provirus in infected cells.

Question 20

In isolating acid-sensitive Escherichia coli using tetracycline-resistant transposons, why is it necessary to screen first for tetracycline resistance and then for acid sensitivity?

Accepted Answer

One screens first for tetracycline resis

Exam 12: Biotechniques and Synthetic Biology

How were GadE and GadX demonstrated to be transcription factors for the gadA gene?

What is synthetic biology? Provide an example of its potential applications.

His₆-fusion-tagged proteins can be recovered from affinity chromatography columns because the histidine tag tightly binds to beads coated with ________ ions. The elution buffer contains ________, which has stronger affinity for these ions.

Noise in synthetic biology is a result of the

For a reporter protein to be synthesized, its gene must be fused in the proper codon reading frame with respect to the

Describe the difference between autoradiography and phosphor imaging.

Potential applications of bacterial oscillator switches include all of the following EXCEPT the

How do operon fusions reveal transcriptional control of a target gene?

Which technique is used to select novel enzymes or peptides produced through directed evolution?

Which kind of phage vectors are most frequently used in phage display technology?

The colorless molecule o-nitrophenyl galactoside (ONPG) can be used as an artificial substrate to assay b-galactosidase in cell extracts of Escherichia coli in which lacZ is used as a reporter gene. Which of the following statements is NOT true with respect to the use of ONPG?

Potential vaccine antigens made by transgenic plants include

The figure below depicts a "kill switch" or suicide module designed to eliminate an organism at a predetermined point, namely once the microorganism's function is accomplished. Explain how this kill switch can be regulated to prevent the untimely death of a microorganism harboring it.

How can a protein be detected on a western blot?

Use the figure below to describe how the CRISPR-Cas9 editing tool is used in eukaryotic cells.

Scientists have taken the genome of the bacterial species ________ and moved it to yeast.

Which probing technique detects DNA immobilized on a membrane using labeled DNA fragments?

CRISPR-Cas9, a prokaryotic defense mechanism, is a powerful emerging tool in biotechnology for all of the following EXCEPT

In isolating acid-sensitive Escherichia coli using tetracycline-resistant transposons, why is it necessary to screen first for tetracycline resistance and then for acid sensitivity?

Microbial Life: Origin and Discovery

Observing the Microbial Cell

Cell Structure and Function

Bacterial Culture, Growth, and Development

Environmental Influences and Control of Microbial Growth

Viruses

Genomes and Chromosomes

Transcription, Translation, and Bioinformatics

Gene Transfer, Mutations, and Genome Evolution

Molecular Regulation

Viral Molecular Biology

Energetics and Catabolism

Electron Flow in Organotrophy, Lithotrophy, and Phototrophy

Biosynthesis

Food and Industrial Microbiology

Origins and Evolution

Bacterial Diversity

Archaeal Diversity

Eukaryotic Diversity

Microbial Ecology

Microbes in Global Elemental Cycles

Human Microbiota and Innate Immunity

The Adaptive Immune Response

Microbial Pathogenesis

Microbial Diseases

Antimicrobial Therapy

Clinical Microbiology and Epidemiology

Filters

His₆-fusion-tagged proteins can be recovered from affinity chromatography columns because the histidine tag tightly binds to beads coated with ions. The elution buffer contains , which has stronger affinity for these ions.