Onward

Question 1

If a person becomes infected after ingesting only a few cells of a pathogen, this organism is said to have a low

Answer 1

infectious dose

Food-borne bacteria, including Escherichia coli, cannot survive the highly acidic environment of the human stomach. An acid-resistant mutant such as E. coli O157:H7 resists environments with pH as low as 2 and therefore even a few cells may be enough to infect the intestinal tract.

Question 2

A protocol includes the following steps: cross-link putative DNA-binding proteins with DNA, shear the DNA, and use an antibody to pull down the protein along with associated DNA. This is a technique called

Answer 2

chromatin immunoprecipitation

Chromatin immunoprecipitation, or ChIP, is a method used to isolate DNA-binding proteins.

Question 3

After performing chromatin immunoprecipitation, you want to determine the DNA sequences pulled down. Your first step is to

Answer 3

amplify the DNA in the presence of fluorescent nucleotides

Prior to exposing the isolated DNA to a DNA microarray, you must first use PCR to amplify the DNA in the presence of fluorescent nucleotides.

Question 4

An application of engineered genetic switchboards is the

Answer 4

design of fail-safe kill mechanisms leading to the timely suicide of genetically modified bacteria

The elimination of genetically modified organisms released outside the laboratory after they have fulfilled their function, but not earlier, can be accomplished by a suicidal mechanism. For example, a crRNA-ccB, which encodes the DNA-damaging protein CcdB, would kill the cell too quickly, unless a modulation mechanism such as an arabinose-induced taRNA, is present.

Question 5

An application of synthetic biology based on bacterial oscillator switches is the

Answer 5

design of bacterial genetic circuits to detect toxins

The interactions between two feedback genetic loops result in a genetic switch that can be toggled depending on the concentration of the inducers. Toxic chemicals, such as arsenic, can be detected by a gfp-fusion system that is activated in the presence of the offending substance.

Question 6

Certain crystallized proteins of Bacillus thuringiensis are used as

Answer 6

insecticides

The Bt toxin, originally produced by Bacillus thuringiensis, is used to kill the larvae of certain crop pests.

Question 7

Gene fusions can provide information about translational control because the

Answer 7

reporter gene must use the ribosome-binding site of the target gene

By using the subject gene’s ribosome-binding site, we can detect whether any factor may affect translation of the subject gene.

Question 8

Genetic switchboards use cis-repressed and trans-activating RNA (crRNA/taRNA) combinations. What type of RNA molecules are cr- and taRNA?

Answer 8

Specially engineered small RNAs

Based on the properties of previously known small RNAs in stabilizing and regulating mRNA translatability, synthetic biologists may design riboswitches (crRNA/taRNA) to control the translation of nearly any target gene.

Question 9

GFP fusions are preferred over LacZ fusions in the study of phenomena such as gene expression in individual cells because

Answer 9

GFP allows the tracking of gene expression in individual cells

the enzymatic activity of LacZ represents the average of a population

LacZ fusions can be used to monitor gene expression by determining enzymatic activity of a mixed population of cells containing LacZ fusions and cells without them. GFP-fusion proteins can be observed in individual cells within a population by microscopy.

Question 10

His6 tagged proteins can be separated from other proteins, based on the affinity of the histidine cluster for

Answer 10

nickel

The His6 tagged proteins can be affinity purified by passing the cell contents through a column containing nickel-coated agarose beads. The tagged protein will stick to the nickel, whereas untagged proteins pass through. A subsequent step can elute the tagged proteins.

Question 11

In fluorescence resonance energy transfer (FRET), a molecule is labeled with a florescent dye and the

Answer 11

energy released by the fluorescent dye is absorbed by a second, nearby molecule

For energy transfer to happen, the second molecule must absorb the energy and the two molecules must be in close proximity.

Question 12

In order to purify a protein by affinity tagging, one can create a fusion between the protein and

Answer 12

His6

His6-tagged proteins can be purified on a nickel column or nickel beads. A fusion with GFP or, β-gal, can elucidate transcriptional activity and is used as a reporter fusion, not to purify a protein. Taq polymerase is a thermostable DNA polymerase used in PCR, not for protein purification.

Question 13

In the electrophoretic mobility shift assay, DNA with a bound protein will run

Answer 13

more slowly through a gel than the DNA without a bound protein

Migration through a gel depends on the molecular weight of the migrating species. The DNA protein complex has a larger molecular weight than the free DNA and thus migrates more slowly.

Question 14

One of the engineering principles used in synthetic biology is

Answer 14

logic gates

A promoter and a gene constitute a genetic logic gate. Bayes theorem is a genetic tool to determine conditional probability. Quantum mechanics is not yet widely used in biology and, broadly defined, materials science studies the microscopic and macroscopic properties of matter.

Question 15

One technique to create random mutations in bacterial genes uses

Answer 15

transposons

Transposons can insert themselves into genes, thus disrupting them and causing mutations.

Question 16

Real-time PCR can give information similar to that given by

Answer 16

northern blots

Although northern blots can be used to quantify specific RNA molecules present in cell extracts, real-time PCR allows us to measure the relative abundance of different RNA species in a sample.

Question 17

Real-time PCR differs from standard PCR in that

Answer 17

real-time PCR is quantitative

Unlike traditional PCR, real-time PCR includes a reporter probe and analyzes samples throughout the experiment. These changes allow real-time PCR to be quantitative and can be used to determine the amount of starting template.

Question 18

The branch of science that is concerned with the development of biological systems that do NOT exist in nature is called

Answer 18

synthetic biology

Biotechnology uses living organisms (wild type or genetically modified) in the manufacture of useful products; this includes directed evolution, in which specific molecular phenotypes are selected, and recombinant DNA, in which natural or synthetic pieces of DNA are joined in vitro. Synthetic biology is biology by design.

Question 19

The yeast two-hybrid assay relies on which of the following techniques?

Answer 19

Fusion proteins

The gene for the yeast GAL4 regulator was split in two functional halves that were then cloned separately. A fusion protein is made from one of the proteins of interest and the GAL4 DNA-binding domain (bait). Another fusion protein is constructed with the second protein of interest and the GAL4 transcriptional activation domain (prey). If the proteins fused to the disjointed domains of GAL4 interact, GAL4 reorganizes, binds to the GAL1 promoter, and starts transcription of the reporter gene lacZ.

Question 20

To determine whether the transcription of a certain gene varies in response to an environmental stimulus, the best option is to

Answer 20

fuse the promoter of the gene to a reporter construct and check reporter activity in the presence and absence of the stimulus

A reporter construct is an appropriate technique to assay transcriptional changes. Both Southern blotting and PCR are methods to detect specific regions of DNA. The DNA for the gene will not change in the presence or absence of the stimulus. Creating a mutant may shed light on the function of the gene but will not give information on transcriptional control.

Question 21

What are BioBricks and what are their uses?

Answer 21

Connectible DNA pieces that can direct the manufacture of novel compounds

DNA segments of known sequence and function have been made publicly available to be used as building materials for engineering new genetic circuits with a wide diversity of applications.

Question 22

What is the purpose of using a fluorescently labeled or enzyme-linked secondary antibody in a western blot experiment?

Answer 22

To detect the presence of the primary antibody on the blot through fluorescence or a color-producing enzymatic reaction

After washing off nonbound antibodies (IgGs), the location of a protein-primary antibody complex is evidenced by the attachment of a secondary IgG that either has a fluorescent tag or is linked to an enzyme that forms a colored product from a colorless substrate.

Question 23

What type of interactions does the yeast two-hybrid assay detect?

Answer 23

Protein-protein

The yeast two-hybrid assay can detect protein to protein binding that might be involved in gene expression control, signal transduction, or other phenomena involving in vivo interactions among proteins.

Question 24

Which enzyme can help determine the DNA sequence directly in contact with a particular DNA-binding protein?

Answer 24

DNase I

DNase I is used in DNA protection assays. The region of DNA bound by a protein is protected from DNase I digestion.

Question 25

Which enzyme is required for primer extension analysis?

Answer 25

Reverse transcriptase

Reverse transcriptase is used to extend the primer back to the 5′ end of the mRNA.

Question 26

Which of following techniques could be useful in determining if a particular protein is present in a cell extract?

Answer 26

Western blot

Western blots can be probed with antibodies specific to a protein of interest.

Question 27

Which of the following elements would be present in a plasmid used for protein expression and affinity purification, but not present in a simple cloning plasmid?

Answer 27

A sequence encoding His6

Nearly all research plasmids contain an origin of replication, an antibiotic resistance gene, and a multiple cloning site. The His6 sequence is unique to plasmids designed for protein purification.

Question 28

Which of the following is a potential disadvantage to producing vaccines in plants?

Answer 28

There may be environmental concerns with genetically modified plants

Some people worry about the escape of genetically modified plants into the wild. Lack of refrigeration is an advantage. Vaccine production in plants can be inexpensive, and a variety of edible plants such as corn and potatoes can be used.

Question 29

Primer extension analysis is a technique to determine the transcriptional start site of a gene?

Answer 29

In primer extension analysis, reverse transcriptase extends a primer that is complementary to a sequence near the 5 end of the RNA being analyzed. The sequence information produced can lead to the determination of the transcriptional start site.

Question 30

Phage display laboratory techniques depend on viruses to study directed evolution of peptides?

Answer 30

In phage display, DNA sequences encoding random peptides are cloned into capsid protein genes of single-stranded DNA phages. Phages displaying peptides with desired properties can be selected.

Question 31

Operon fusions reflect transcriptional control of the subject gene, whereas gene fusions reflect both transcriptional and translational control.

Answer 31

Both types of fusions give information about transcriptional control. Since operon fusions have their own ribosome-binding sites but gene fusions do not, gene fusions also give information about translational control.

Question 32

Which of the following techniques is best suited to visualize the subcellular localization of a protein?

Answer 32

GFP-fusion proteins

Green fluorescent protein fusions can light up a protein of interest so the protein’s cellular localization can be observed by microscopy. Western blotting can determine the presence of a protein in an extract, but not its cellular localization.

Question 33

Which of the following techniques is most likely to use fluorescence resonance energy transfer (FRET)?

Answer 33

Real-time PCR

Real-time PCR employs FRET to monitor DNA synthesis as it occurs.

Question 34

Which of the following techniques relies on nucleic acid hybridization to detect an mRNA of interest?

Answer 34

Northern blot

Northern and Southern blotting use nucleic acid hybridization to detect RNA and DNA, respectively. In western blotting, antibodies are used to detect proteins. In denaturing polyacrylamide gel electrophoresis (SDS-PAGE), proteins are separated based on their molecular weight

Question 35

You have isolated a bacterial transcription factor using ChIP. What is the next step in order to find out all the sites in the genome that can be bound by this particular protein?

Answer 35

DNA co-immunoprecipitated with the protein is amplified and identified on a microarray (ChIP-on-chip)

Each spot on the microarray chip contains a small segment of the genome of the bacterium under study, indicating which specific genes may be controlled by the transcription factor