CRISPR and their applications to genetic engineering

Question 1

What does CRISPR stand for and what does it do

Answer 1

CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats and they are regions in prokaryotic genomes that provide a defense against viral infection.

Question 2

How is CRISPR in prokaryotes analogous to vertebrates’ immune system

Answer 2

CRISPR in prokaryotes is loosely analogous to vertebrates’ immune system in the way that they can be created in response to a viral challenge. In addition, they are inherited ( an example of inheritance of acquired characteristics). The mechanism is completely different.

Question 3

Describe the components of CRISPR loci

Answer 3

CRISPR loci contain copies of repeat sequences that are separated by spacers. They are linked to genes encoding CRISPR-associated (Cas) proteins, which function as nucleases. They store memory of past infections. Upon subsequent reinfection by the virus, the cell detects DNA matching a CRISPR sequence, triggering the Cas nuclease to cleave it.

Question 4

Name another organism that contains a CRISPR-like system

Answer 4

Large mimiviruses also contain CRISPR-like systems. This is a defense mechanism against ‘virophages’ that infect the virus.

Question 5

Discuss the mechanism behind CRISPR

Answer 5

There are 3 stages- adaptation, expression, and interference. Adaptation- when a phage infects a bacterium, assuming the cell survives, regions of the phage DNA are clipped out, replicated then integrated into a new CRISPR locus, separated by spacers. Expression- transcription of the new regions produces CRISPR-RNAs (cRNAs) that bind to Cas proteins. Interference- the Cas proteins use the bound RNA as a kind of probe. When a match is detected to a region of DNA from an invading virus, the Cas protein clips the viral DNA. This degradation of the viral DNA is a defense mechanism of the cell.

Question 6

In the application of CRISPR/Cas techniques, there is gene knockout and gene editing. Describe the differences between them

Answer 6

In both gene knockout and gene editing, there is a synthetic guide RNA targeting a genomic region of interest