MCB4203 - Module 7 Exam Latest
Update
Isolation and purification of bacterial product (biochemical methods) leads to
demonstration of toxicity and/or disease symptoms - Answer What was the "old way" of
identifying virulence factors?
1. More rapid discovery of virulence genes
2. Discovery of virulence traits not previously suspected - Answer What are two things
advances in molecular biology (past 25 years) led to breakthroughs in?
Second wave revolution - Answer What did whole-genome sequencing, microarrays,
automation, and advanced bioinformatics lead to?
1. Growth of bacterial culture in vitro
2. Fractionation of culture supernatant
3. Screen fractions for virulence phenotype
4. Purify factor from positive fractions
5. Retest purified factor in vivo, should reproduce some or all disease symptoms
6. Reverse genetics, recombinant protein expression/purification, antigenicity - Answer
What are the six steps of the typical "biochemical approach"?
Affinity chromatography - Answer What uses a bound receptor or ligand and an eluent
with free ligand or a receptor for the protein of interest?
Rabbit ileal loop assay - Answer What uses the intestine to perform a typical
biochemical approach to study the effect of cholera toxin; loop lengths and amount of
fluid accumulated in each loop are measured?
1. Virulence factor is produced when bacteria is cultured in vitro
2. Toxin always results in host cell damage - Answer What are the two assumptions that
are used for virulence factors to be identified by the biochemical method?
Slow (years to decades) - Answer Is the typical "biochemical approach" fast or slow?
Bacterial toxins (host-cell damaging proteins)
Secreted enzymes
LPS - Answer What three things does the typical "biochemical approach" primarily work
on?
, 1. Chemical or UV mutagenesis
2. Expression of virulence factors in avirulent strain - recombinant genes
3. Transposons
4. Reporter fusion - Answer What four things are apart of the molecular "genetics
approach"?
Chemical/UV mutagenesis - Answer What checks mutants for in vivo virulence?
Useful if pathogen is difficult to manipulate genetically - Answer What is the advantage
of chemical/UV mutagenesis?
Difficult to identify mutations (not marked) possibly more than one "hit" per strain -
Answer What is the disadvantage of chemical/UV mutagenesis?
Expression of virulence factor in an avirulent strain - Answer What is best applied to
closely-related species, can be time-consuming/tedious, can only analyze small
segments of genome?
Gram-negative bacteria - Answer When identifying virulence genes by cloning and
expressing them in avirulent bacterial strains, which bacteria clones fragments of
genomic DNA from an invasive Salmonella strain into a laboratory strain of E. coli?
Gram-positive bacteria - Answer When identifying virulence genes by cloning and
expressing them in avirulent bacterial strains, which bacteria clones the gene for the
hemolysin listeriolysin O from Listeria monocytogenes into a laboratory strain of
Bacillus subtilis?
Transposon - Answer What is a segment of bacterial DNA that can be translocated as a
whole between chromosomal, phage, and plasmid DNA in the absence of a
complementary sequence in the host DNA?
Contain inverted repeats at their ends
Encode enzymes required for transposition
Often carry antibiotic resistance genes
Can be used as a molecular tool for whole-genome and single-gene studies in bacteria
Used to introduce a piece of foreign DNA - Answer What are 5 characteristics of
transposons?
Transposon library - Answer What contains a group of bacterial mutants that have
transposon insertions in all non-essential genes?
Target site - Answer Transposase binds to both ends of the transposon and a sequence
of DNA that makes up the ______________.
Update
Isolation and purification of bacterial product (biochemical methods) leads to
demonstration of toxicity and/or disease symptoms - Answer What was the "old way" of
identifying virulence factors?
1. More rapid discovery of virulence genes
2. Discovery of virulence traits not previously suspected - Answer What are two things
advances in molecular biology (past 25 years) led to breakthroughs in?
Second wave revolution - Answer What did whole-genome sequencing, microarrays,
automation, and advanced bioinformatics lead to?
1. Growth of bacterial culture in vitro
2. Fractionation of culture supernatant
3. Screen fractions for virulence phenotype
4. Purify factor from positive fractions
5. Retest purified factor in vivo, should reproduce some or all disease symptoms
6. Reverse genetics, recombinant protein expression/purification, antigenicity - Answer
What are the six steps of the typical "biochemical approach"?
Affinity chromatography - Answer What uses a bound receptor or ligand and an eluent
with free ligand or a receptor for the protein of interest?
Rabbit ileal loop assay - Answer What uses the intestine to perform a typical
biochemical approach to study the effect of cholera toxin; loop lengths and amount of
fluid accumulated in each loop are measured?
1. Virulence factor is produced when bacteria is cultured in vitro
2. Toxin always results in host cell damage - Answer What are the two assumptions that
are used for virulence factors to be identified by the biochemical method?
Slow (years to decades) - Answer Is the typical "biochemical approach" fast or slow?
Bacterial toxins (host-cell damaging proteins)
Secreted enzymes
LPS - Answer What three things does the typical "biochemical approach" primarily work
on?
, 1. Chemical or UV mutagenesis
2. Expression of virulence factors in avirulent strain - recombinant genes
3. Transposons
4. Reporter fusion - Answer What four things are apart of the molecular "genetics
approach"?
Chemical/UV mutagenesis - Answer What checks mutants for in vivo virulence?
Useful if pathogen is difficult to manipulate genetically - Answer What is the advantage
of chemical/UV mutagenesis?
Difficult to identify mutations (not marked) possibly more than one "hit" per strain -
Answer What is the disadvantage of chemical/UV mutagenesis?
Expression of virulence factor in an avirulent strain - Answer What is best applied to
closely-related species, can be time-consuming/tedious, can only analyze small
segments of genome?
Gram-negative bacteria - Answer When identifying virulence genes by cloning and
expressing them in avirulent bacterial strains, which bacteria clones fragments of
genomic DNA from an invasive Salmonella strain into a laboratory strain of E. coli?
Gram-positive bacteria - Answer When identifying virulence genes by cloning and
expressing them in avirulent bacterial strains, which bacteria clones the gene for the
hemolysin listeriolysin O from Listeria monocytogenes into a laboratory strain of
Bacillus subtilis?
Transposon - Answer What is a segment of bacterial DNA that can be translocated as a
whole between chromosomal, phage, and plasmid DNA in the absence of a
complementary sequence in the host DNA?
Contain inverted repeats at their ends
Encode enzymes required for transposition
Often carry antibiotic resistance genes
Can be used as a molecular tool for whole-genome and single-gene studies in bacteria
Used to introduce a piece of foreign DNA - Answer What are 5 characteristics of
transposons?
Transposon library - Answer What contains a group of bacterial mutants that have
transposon insertions in all non-essential genes?
Target site - Answer Transposase binds to both ends of the transposon and a sequence
of DNA that makes up the ______________.
