MCB5205 Final Exam With
Complete Solution
Affinity tags - ANSWER Attach antibody to bead, wash with 2 different
buffers
Specific Ligands - ANSWER Attach ligand to bead wash with buffer
Metal-Chelation - ANSWER Tagged and untagged proteins bound to beads,
wash with buffer, elute with imidazole
Rabbit Ileal loop - ANSWER Studying effect of cholera
Rabbit ideal loops inoculated with saline
Loop lengths and fluid amount measured
Advantages and disadvantages of chemical UV mutagenesis - ANSWER
Advantage - useful if pathogen is difficult to manipulate genetically
Disadvantage - difficult to identify mutations
How to express virulence in non-virulent strain? - ANSWER Recombinant
genes
Introduce cloned DNA segments from another strain, retrieve cloned DNA,
sequence + characterize cloned DNA set
Importance of transposons (jumping gene) - ANSWER Can be translated
between chromosomal, phage, plasmid DNA in absence of complementary
sequence
,In host DNA
Used to introduce piece of foreign DNA
Transposon mutagenesis - ANSWER Carry transcriptional terminators
Identity non-essential genes
Need complementation to identify non- essential genes
Complementation - ANSWER WT copy of Transposon disrupted gene
re-introduced at second site
Used to restore gene exp
Can distinguish polar effects on downstream genes in operons
Reporters - ANSWER Enzyme whose activity is easy to measure/detect
Reporter fusion - ANSWER Links regulatory circuit of virulence gene to
reporter gene
Biophotonic imaging - ANSWER Visualize bacteria/host molecules during
infection
Laser capture microscopic dissection ( LCM) - ANSWER can be used to extract
specific/localized regions of interest from tissue
RNA microarray/RNA seq - ANSWER can be used to compare host gene
expression changes in response to infection
Gives into about whole cell during infection
RNAi - ANSWER Used to turn off specific host genes
Proteomics - ANSWER Identifies changes in protein /protein properties
, Knockout mice - ANSWER Deletion/inactivation of gene
Gene-edited mice - ANSWER Target gene edited by CRISPR
Conditional knockout mice - ANSWER Target gene inactivated in specific cell
type
Gene replacement mice - ANSWER Gene replaced wl gene from another
animal
Nude mice - ANSWER Defective in fox-1 gene
Transgenic mice - ANSWER Defective in specific immune cells
Gnotobiotic mice - ANSWER Born aseptic, no resident Microbiota
Specific pathogen free - ANSWER Free of specific pathogens, exposed to
other Microbiota
How was shigella virulence plasmid found? - ANSWER Koch's postulates
Loss of plasmid associated with T to O transition, loss of virulence in S sonnei
T3SS in shigella - ANSWER To secrete proteins
Encoded by Mxi and Spa
Effectors IpaA-D
Target host cell membrane, form pores
Target regulatory/structural host components
Actin tail - ANSWER Allows for propulsion into other cells
Shigella and Listeria
Antibiotic resistance in shigella - ANSWER Transferred by R-factor
Complete Solution
Affinity tags - ANSWER Attach antibody to bead, wash with 2 different
buffers
Specific Ligands - ANSWER Attach ligand to bead wash with buffer
Metal-Chelation - ANSWER Tagged and untagged proteins bound to beads,
wash with buffer, elute with imidazole
Rabbit Ileal loop - ANSWER Studying effect of cholera
Rabbit ideal loops inoculated with saline
Loop lengths and fluid amount measured
Advantages and disadvantages of chemical UV mutagenesis - ANSWER
Advantage - useful if pathogen is difficult to manipulate genetically
Disadvantage - difficult to identify mutations
How to express virulence in non-virulent strain? - ANSWER Recombinant
genes
Introduce cloned DNA segments from another strain, retrieve cloned DNA,
sequence + characterize cloned DNA set
Importance of transposons (jumping gene) - ANSWER Can be translated
between chromosomal, phage, plasmid DNA in absence of complementary
sequence
,In host DNA
Used to introduce piece of foreign DNA
Transposon mutagenesis - ANSWER Carry transcriptional terminators
Identity non-essential genes
Need complementation to identify non- essential genes
Complementation - ANSWER WT copy of Transposon disrupted gene
re-introduced at second site
Used to restore gene exp
Can distinguish polar effects on downstream genes in operons
Reporters - ANSWER Enzyme whose activity is easy to measure/detect
Reporter fusion - ANSWER Links regulatory circuit of virulence gene to
reporter gene
Biophotonic imaging - ANSWER Visualize bacteria/host molecules during
infection
Laser capture microscopic dissection ( LCM) - ANSWER can be used to extract
specific/localized regions of interest from tissue
RNA microarray/RNA seq - ANSWER can be used to compare host gene
expression changes in response to infection
Gives into about whole cell during infection
RNAi - ANSWER Used to turn off specific host genes
Proteomics - ANSWER Identifies changes in protein /protein properties
, Knockout mice - ANSWER Deletion/inactivation of gene
Gene-edited mice - ANSWER Target gene edited by CRISPR
Conditional knockout mice - ANSWER Target gene inactivated in specific cell
type
Gene replacement mice - ANSWER Gene replaced wl gene from another
animal
Nude mice - ANSWER Defective in fox-1 gene
Transgenic mice - ANSWER Defective in specific immune cells
Gnotobiotic mice - ANSWER Born aseptic, no resident Microbiota
Specific pathogen free - ANSWER Free of specific pathogens, exposed to
other Microbiota
How was shigella virulence plasmid found? - ANSWER Koch's postulates
Loss of plasmid associated with T to O transition, loss of virulence in S sonnei
T3SS in shigella - ANSWER To secrete proteins
Encoded by Mxi and Spa
Effectors IpaA-D
Target host cell membrane, form pores
Target regulatory/structural host components
Actin tail - ANSWER Allows for propulsion into other cells
Shigella and Listeria
Antibiotic resistance in shigella - ANSWER Transferred by R-factor
