CRISPR - MT3 - Part 2

Question 1

The bacterial CRISPR system has been modified to do what?

Answer 1

To make genomic changes efficient

Question 2

sgRNA

Answer 2

Single guide RNA

Question 3

What is sgRNA?

Answer 3

Is an engineered molecule that combines the functions of the crRNA and its assistant, the tracrRNA

Question 4

What can CRISPR be used for? (2)

Answer 4

1. Used to KO a gene you dont want (LOF)
   - can be used anywhere you want to insert something into the cell
2. Change the function of a gene (GOF) on the basis of a donor template
   - this sequence only works if it is next to the PAM sequence
   - cas9 mediated cleavage of target DNA, naturally in a virus and as a genome editing tool

Question 5

The system that repairs the break does not restore what?

Answer 5

Original strand

Question 6

What can CRISPR cas9 be used to do? (2)

Answer 6

1. KO a gene

2. Edit it according to the template

Question 7

What does D163 do?

Answer 7

Plays an important role in the immune system, but also acts as a receptor for the porcine respiratory and reproductive syndrome virus
- cant enter unless invited in

Question 8

PRRSV

Answer 8

Porcine respiratory and reproductive syndrome virus

Question 9

What is the gene CD163 modified by? And what does it then do?

Answer 9

1. CRISPR-cas9

2. Allows it to eliminate with PRRSV binding sequence of CD163 without impairing its immune function

Question 10

Where can you find the gene ARG058?

Answer 10

In corn

Question 11

Whats does ARG058 do?

Answer 11

It improves drought resistance, but has a weak promoter
- a stronger promoter was introduced by CRISPR-cas9, resulting in better drought resistance and high crop yield under drought conditions

Question 12

Promoter

Answer 12

DNA sequences that define where transcription of a gene by RNA polymerase begins

Question 13

Where is a promoter typically located?

Answer 13

Directly upstream or at the 5’ end of the transcription initiation site

Question 14

What can the targeting ability of sgRNA be used for? (2)

Answer 14

1. To turn genes on and off, to add epigenetic markers such as methyl groups
2. To localize a given sequence in a cell

Question 15

What do CRISPR functions depend on?

Answer 15

Cas9 nuclease activity being inactivated

- dead cas9 (dcas9)

Question 16

What are the 4 categories that are/will be looked at as soon as possible?

Answer 16

1. Editing cultured human cells for research purposes
   - cultured human cells can divid forever
2. Editing somatic cells for medical research
   - such as treating diseases
3. Editing germ-line cells or embryos to prevent transmission of diseases
4. Editing cells (somatic of germ-line) for reasons fo enhancement, not because of disease