BioSc221/325 Exam 2 Name ______________________________
Multiple choice.
(1 point each) Choose the one best answer to each of the following questions.
____ What is the function of dideoxynucleotides in DNA sequencing?
A. Dideoxynucleotides provide a 3' hydroxyl residue for chain elongation.
B. Dideoxynucleotides cause chain termination leading to DNA fragments of varying length.
C. Dideoxynucleotides provide a radioactive label for detecting bands on a sequencing gel.
D. Dideoxynucleotides function as a primer to initiate chain elongation.
____ Which of the following mechanisms for horizontal gene transfer in nature would not work in a strain that is deficient in homologous recombination?
A. conjugation
B. lysogeny
C. transformation
____ Genes encoding the enzymes required for synthesis of the amino acid tyrosine are likely to be controlled by what type of repressor?
A. inducible
B. repressible
C. constitutive
____ A plasmid vector with a single origin of replication that is capable of replicating in bacteria from more that one genera are called a
A. shuttle vector
B. suicide vector
C. cosmid vector
D. broad host range vecor
____ Bacterial protein involved in homologous recombination and is required for repair of UV damage to DNA.
A. helicase
B. topoisomerase
C. repairase
D. RecA
____ The origin of replication of a plasmid cloning vector can affect
A. compatibility with other cloning vectors
B. the copy number of the plasmid in the host bacterium
C. the direction of replication
D. all of the above
____ UV light is lethal to bacterial cells due to formation of
A. pyrimidine dimers which, when they are removed, can result in double stranded breaks in the DNA.
B. apurinic sites which are easily broken.
C. double stranded breaks resulting from direct cleavage by the UV radiation.
D. creation of multiple nonsense codons that disrupt translation.
____ If a suicide vector is used to introduce a cloned gene into a new host bacterium, how is the cloned gene maintained in the new host?
A. homologous recombination with the host cell chromosome
B. replication of the suicide vector
C. conjugative transfer using rolling circle replication
D. by becoming lysogenic

____ Protein involved in transcription initiation by aiding in promotor recognition.
A. core enzyme
B. stem loop
C. rho
D. sigma
____ Genes that are expressed at the same level at all times are said to be
A. constitutive
B. inducible
C. repressible
D. activated
____ Most prokaryotic promoters have a two-part consensus sequence that consists of a
A. promoter and terminator
B. promoter and operator
C. activator and operator
D. –10 and –35 region
____ The primary sigma factor in Escherichia coli is called
A.

70
B.

18
C.

54
D.

P
____ Several genes that are transcribed in the same direction and share the same promoter are called a(n)
A. operator
B. operon
C. polylinker
D. polypromoter
____ Most plasmids inside a bacterial cell can be found in what condition?
A. covalently closed circles that are supercoiled
B. covalently closed circles that are relaxed
C. linear strands
D. all of the above
____ In two-component regulatory systems a signal is relayed to the regulator protein by
A. phosphorylation
B. adenylation
C. autoinducer
D. a peptide
____ Promoter strength is determined by
A. the DNA sequence of the promoter region
B. the number of operators
C. how close the promoter is to the start of transcription
D. the DNA sequence of the ribosome binding site
____ Reporter gene fusions are used to study gene expression because
A. not all enzyme activities are easily measured
B. feedback inhibition of the enzyme being studied may interfere with activity levels
C. post-transcriptional or post-translational factors associated with the gene being studied may interfere with activity levels
D. all of the above

____ The antibiotic to the right is a(n)
A. aminoglycoside
B.

-lactam
C. chloramphenicol
D. macrolide
E. tetracycline
____ The antibiotic to the right is a(n)
A. aminoglycoside
B.

-lactam
C. chloramphenicol
D. macrolide
E. tetracycline
____ If you wan to study transcription of the pcl gene, which codes for the protein Pcl, you should construct a transcriptional fusion to the lacZ gene. This involves (RBS = ribosome binding site)
A. cloning the pcl promoter upstream of a lacZ gene which lacks a promoter but still contains a native lacZ RBS.
B. cloning the pcl promoter and RBS upstream of the lacZ gene that lacks the native lacZ RBS.
C. cloning the pcl promoter downstream of a lacZ gene which lacks a promoter but still contains a native lacZ RBS.
D. cloning the pcl promoter and RBS downstream of the lacZ gene that lacks the native lacZ RBS.
____ A bacterial strain in which an F plasmid has integrated into the chromosome allowing easy transfer of chromosomal DNA to a recipient strain is called a(n)
A. conjugative transposon
B. conjugative chromosome
C. R factor
D. HFR
___ Quorum sensing systems use ___ to determine cell density.
A. cell-cell contact
B. autoinducer
C. luminescence
D. phosphotransferase

Matching.
(1 point each) Match the definition in the left column with the appropriate term in the right column. Note, not all terms will be used.
____ Type of autoinducer found in Gram-negative bacteria.
____ Plasmid with more than one origin of replication for use in organisms from two different
kingdoms
____ Stem loop structure that forms at the end of a mRNA.
____ The membrane component of a two-component regulatory system.
____ Bacteriophages that can transfer random regions of the host chromosome
____ Antibiotic resistance gene flanked by two insertion sequences.
____ Mechanism by which Streptococcus pneumoniae can acquire penicillin resistance genes.
____
 ’ subunits of RNA polymerase.
____ Endospores contain a high concentration of ___ which probably contributes to resistance.
____ DNA sequence at which repressors bind to turn off gene expression.
Short answer. (1 point each)
A.
Activator
B.
Broad-host range vector
C.
Calcium dipicolinate
D.
Core enzyme
E.
Diaminopimelicacid
F.
Generalized transducing phage
G.
Homologous recombination
H.
Homoserine lactone
I.
Operator
J.
Promoter
K.
Reduction
L.
Rolling circle replication
M.
Selectable marker
N.
Sensor
O.
Shuttle vector
P.
Specialized transducing phage
Q.
Telomere
R.
Terminator
S.
Transposon
Irradiated food is generally considered sterile and at least has a very long shelf life. However, Deinococcus radiodurans was isolated from irradiated meat. How did it manage to survive this "sterilization" process?
What is the definition of a cloning vector?
Name a genus of bacteria that is capable of producing endospores.
Diphtheria toxin is made up of two proteins, A and B, which are present in a ratio of 1:5 respectively. The genes that code for these proteins are part of an operon. How can two genes that belong to a single operon be expressed at such dramatically different levels?
A strain of E. coli was recently isolated from a hospital patient and found to be resistant to the antibiotic chloramphenical.
What would be a likely mechanism for this resistance phenotype?

BvgR is the regulatory protein component of a two-component regulatory system found in E. coli . BvgR functions as a repressor of some genes and as an activator of others. Describe the likely location of the DNA binding site for BvgR when it is functioning as an:
Activator
Repressor
Mike Robe, ace microbiologist at the Centers for Disease Control, has been studying a new, highly virulent strain of
Escherichia coli that has been responsible for several outbreaks in south central Missouri. He has identified a cluster of genes that seems to be responsible for the virulent nature of this new isolate. One significant discovery he has made is that the G+C ratio of this virulence gene cluster is much lower than the G+C content of the rest of the E. coli genome, which is now known.
What is the significance of this observation?
Bea Subtilis, head microbiologist at Phelps County Regional Medical Center, noticed an increased frequency of multi-drug resistant strains of Salmonella in patients with food poisoning. Using her finely honed epidemiological skills, she determined that the source of the outbreak was a community with a large number of hog farms in the surrounding area. She blamed the outbreak on the commercial hog farms. Why?
When cloning genes using E. coli it is preferable to use a strain of E. coli that does not make the enzyme RecA ( recA
-
). Why do you think a recA
-
strain is better than a recA + strain for gene cloning?
What is the difference between a transposon and an insertion sequence?
Before a conjugative transposon can be transferred to a recipient strain it must excise itself from the chromosome and form a double-stranded circular molecule that is similar to a conjugative plasmid. Why is this necessary? (hint: what is the mechanism for transfer of the conjugative transposon?)
Biotech companies are constantly developing new cloning plasmids with various "Bells and Whistles" that make screening or selecting for recombinant plasmids easier. Give one example of such an added feature.

Short Essay Questions.
Please answer the following three short essay questions (5 points each)
Describe the process of endospore formation. Explain why endospores are highly resistant to dessication and chemical damage.
In your new job as a microbiologist for MegaDrugs Pharmaceuticals, Inc. you are assigned the task of characterizing a collection of mutant strains of Erwinia caratovora isolated by your predecessor. Looking through your predecessors wellmaintained lab notes you read that she had used chemical mutagenesis to generate mutations and then screened for strains that no longer made active pectinase. Your job is to determine the reason for loss of enzyme activity in each strain. What are the possible explanations for the loss of enzyme activity?

Streptococcus pneumoniae is described as naturally competent for uptake of "naked" DNA. Explain what this means. Discuss all the possible mechanisms - natural and artificial - for uptake of "naked" DNA by bacteria.
Bonus. (3 points possible) In just the past few years, we have seen the determination of the entire genome sequences of dozens of bacterial species. Describe one example of how this information is being used for the benefit of mankind.