Lecture 10: CRISPR

Why do bacteria have CRISPR?

To be able to remember which viruses they have been infected with

To cut apart virus DNA thereby stopping the virus infection

How does CRISPR edit DNA?

CRISPR cuts the DNA and the cell makes errors when it tries to fix the broken DNA

CRISPR cuts the DNA and the cell can use a similar DNA sequence as a template to try to fix the DNA

After Cas9 cuts the target DNA, cellular enzymes attempt to repair the break. Which of the following repair mechanisms involves "editing" of the gene with insertion of a corrected sequence?

Homology Directed Repair

What is the role of PAM (Protospacer Adjacent Motif) in CRISPR-Cas9 system?

PAM is a recognition site for the Cas9 enzyme in the CRISPR-Cas9 system.

In the CRISPR Out of the Blue lab, gene editing will occur in the bacteria if

The bacteria is exposed to arabinose

The bacteria is transformed with the Donor Guide plasmid (pDG)

We have been modifying DNA sequences for many years - ever since restriction enzymes were discovered in the early 1970's. CRISPR and restriction enzymes are essentially the same because they can both be used to cut DNA and can be used inside of living cells.

FALSE - Restriction enzymes cannot be used inside of living cells

What is the function of Cas1/Cas2 in bacterial adaptive immunity?

Removes segment of foreign DNA

What does CRISPR allow scientists to do?

Deleting, Adding, or Changing Sections of DNA

What does the acronym CRISPR stand for?

Clustered Regularly-Interspaced Short Palindromic Repeats

Which enzyme is responsible for cutting the target DNA in CRISPR-Cas9 system?

Cas9

A PAM is

a 3 bp sequence that adjacent to viral DNA in a CRISPR array

The Cas enzymes (Cas1, 2, & 9) are all examples of...

endonucleases

What is the goal of the CRISPR Out of the Blue lab?

To edit the lacZ gene and prevent the metabolizing of X-gal

What is the function of guide RNA in CRISPR-Cas9 system?

Guide RNA guides the Cas9 enzyme to the target DNA sequence.

Explain the role of the guide RNA (gRNA) in the CRISPR-Cas9 mechanism

It binds to the target DNA sequence and guides the Cas9 enzyme to the correct location.

CRISPR-Cas9 was discovered in which of the following organisms?

bacteria

Which of the following could be an example of a PAM?

AGG

Which of the following is NOT a component of the CRISPR Cas9 system?

DNA polymerase

What term describes the segments of viral DNA inserted into the CRISPR region of the bacterial genome?

spacers

After Cas9 cuts the target DNA, cellular enzymes attempt to repair the break. Which of the following types of repair mechanisms creates a mutation and inactivates the gene.

Non-Homologous End Joining

What is the role of Cas9?

Cut DNA at specific locations.