BCH 5413 EXAM ONE QUESTIONS
WITH VERIFIED ANSWERS
Where in the genome are loxP sites located in a Cre/Lox model? - Answer-On either
side of the gene that is to be knocked out
In which organism was the CRISPR system discovered? - Answer-e. coli
In the immunization phase, what happens to the foreign viral DNA? - Answer-Cleaved
into pieces by cas complex and the fragments are called spacers. Some of the viral
fragments will become a part of the sequence.
What RNAs are present in the Cas9 complex? And what is their role? - Answer-
tracrRNA: binds to repeat sequences
What happens in a new infection from a previously recognized viral DNA? - Answer-
Complementary basepair to spacer region and if PAM sequence is present than Cas9
will act as a nuclease and the infection can't go on.
What is the importance of PAM sequence in CRISPR Cas9 technology? - Answer-The
presence of PAM allows Cas9 to function as a nuclease
What are the 2 possible repair mechanisms of a double strand break? Which one of the
methods would you prefer if you wish to make a three nucleotide change in your target
gene using CRISPR Cas9? - Answer-Non-homologius end joining
Homology directed repair: you would want to use this one because you can inject new
donor DNA
In the example of mdx gene mutation given in lecture, why is the successful CRISPR
Cas9 mutation detected as a band of the same size as wild type in the undigested PCR
panel and as smaller band in the TseI digested PCR panel? (panel E) - Answer-The
presence of the TSE fragment indicates a cut in the DNA. There were no cuts in the non
Tsel.
In the Zinc Finger approach to gene editing, what would be the equivalent of the
guideRNA from CRISPR? Why? (explain the function of each) - Answer-Zinc finger
domains recognize DNA and folk1 enzyme will cleave DNA
In the example given, what is the importance of the CCR5 gene? Briefly describe how
this technology is helpful in controlling the replication of HIV-1. - Answer-CCR5 is the
protein on the surface of the T cells that HIV binds to and gets itself into the cell where it
will replicate and eventually burst the T cell and it will go off to infect others. This
technology is helpful because they can gene edit persons infected with HIV to not have
CCR5 so that then their HIV-1 infection cannot progress any further.
What are the two components of the prokaryotic RNA polymerase holoenzyme? What is
the function of each? - Answer-Core enzyme: contains catalytic activity, synthesizes
RNA randomly on dsDNA templates
, Sigma factor: confer promoter binding and specificity, not required for elongation
Briefly describe how the DNA footprinting experiment works. What regions of the
promoter were shown to be important for RNAP binding based upon the DNA footprint?
WITH VERIFIED ANSWERS
Where in the genome are loxP sites located in a Cre/Lox model? - Answer-On either
side of the gene that is to be knocked out
In which organism was the CRISPR system discovered? - Answer-e. coli
In the immunization phase, what happens to the foreign viral DNA? - Answer-Cleaved
into pieces by cas complex and the fragments are called spacers. Some of the viral
fragments will become a part of the sequence.
What RNAs are present in the Cas9 complex? And what is their role? - Answer-
tracrRNA: binds to repeat sequences
What happens in a new infection from a previously recognized viral DNA? - Answer-
Complementary basepair to spacer region and if PAM sequence is present than Cas9
will act as a nuclease and the infection can't go on.
What is the importance of PAM sequence in CRISPR Cas9 technology? - Answer-The
presence of PAM allows Cas9 to function as a nuclease
What are the 2 possible repair mechanisms of a double strand break? Which one of the
methods would you prefer if you wish to make a three nucleotide change in your target
gene using CRISPR Cas9? - Answer-Non-homologius end joining
Homology directed repair: you would want to use this one because you can inject new
donor DNA
In the example of mdx gene mutation given in lecture, why is the successful CRISPR
Cas9 mutation detected as a band of the same size as wild type in the undigested PCR
panel and as smaller band in the TseI digested PCR panel? (panel E) - Answer-The
presence of the TSE fragment indicates a cut in the DNA. There were no cuts in the non
Tsel.
In the Zinc Finger approach to gene editing, what would be the equivalent of the
guideRNA from CRISPR? Why? (explain the function of each) - Answer-Zinc finger
domains recognize DNA and folk1 enzyme will cleave DNA
In the example given, what is the importance of the CCR5 gene? Briefly describe how
this technology is helpful in controlling the replication of HIV-1. - Answer-CCR5 is the
protein on the surface of the T cells that HIV binds to and gets itself into the cell where it
will replicate and eventually burst the T cell and it will go off to infect others. This
technology is helpful because they can gene edit persons infected with HIV to not have
CCR5 so that then their HIV-1 infection cannot progress any further.
What are the two components of the prokaryotic RNA polymerase holoenzyme? What is
the function of each? - Answer-Core enzyme: contains catalytic activity, synthesizes
RNA randomly on dsDNA templates
, Sigma factor: confer promoter binding and specificity, not required for elongation
Briefly describe how the DNA footprinting experiment works. What regions of the
promoter were shown to be important for RNAP binding based upon the DNA footprint?
