FN - CRISPR Flashcards
What does CRISPR stand for?
CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeats
What is the CRISPR-Cas system? (4)
- It is a bacterial defence system that protects bacteria from viruses
- CRISPR arrays are clusters of short, repeated DNA sequences that are interspaced with spacer sequences
- Spacer sequences are derived from the DNA of viruses that have infected the bacterium
- Cas genes encode proteins that help the CRISPR system to target and destroy viral DNA
What do Cas proteins do? (3)
These proteins are encoded by the Cas genes and are responsible for various tasks within the CRISPR-Cas system, including:
- unwinding target DNA
- Snipping invader DNA
- Destroying foreign genetic elements
What is the difference between Class 1 and Class 2 CRISPR-Cas systems?
Class 1 CRISPR-Cas systems are more complex and have multi-subunit effector complexes
- RNA guided
- DNA binding and cleavage
Class 2 CRISPR-Cas systems are simpler and have single protein effector modules
- Dual RNA-guided
- DNA cleavage
Name the types and subtypes of CRISPR-Cas systems
Class 1
- Type I: This type includes subtypes I-A, I-B, I-C, I-E, and I-F [1].
- Type III: This type includes subtypes III-A and III-B [1].
- Type IV: This type is shown but has no subtypes listed [1].
Class 2
- Type II: This type is the most common type and includes subtypes II-A, II-B, and II-C [1].
- Type V: This type includes subtypes V-A, V-B, V-C, V-D, V-E, V-U1, V-U2, V-US, and V-O3 [1].
- Type VI: This type includes subtypes VI-A, VI-B1, VI-B2, and VI-C [1]
What is CUT-PCR?
CUT-PCR stands for CRISPR-mediated Ultrasensitive PCR.
What are the key components of the Type I-E CRISPR-Cas system? (2)
This is a subtype of the CRISPR-Cas systems found in bacteria and archaea that provide defense against viruses and plasmids
- Cascade (CasABCDE) is involved in recognising target sequences
- Cas3 cleaves the target DNA
Describe the crRNA biogenesis process (3)
- CRISPR forms pre-crRNA
- CasE complex: involved in crRNA maturation
- Pre-crRNA processing/maturation: the longer precursor RNA molecule is cleaved into individual crRNAs
Summarize the 3 stages of CRISPR-Cas immunity
i) ADAPTATION
1) New virus enters
2) Cas-associated proteins acquire spacer
3) CRISPR array expanded
ii) EXPRESSION AND crRNA MATURATION
4) Cas proteins formed and crRNA matured
5) Cascade formed
iii) INTERFERENCE
6) Known virus enters
7) Cascade recognises virus DNA and recruits the Cas3 nuclease-helicase
8) Virus DNA degraded
How can CRISPR-Cas be dangerous for the host? (2)
- Can sometimes introduce unintended mutations at sites other than the intended target sequence
- The Cas9 protein can trigger an immune response
Describe the process of invader DNA recognition and autoimmunity prevention. (3)
1) Scan for PAM (Protospacer Adjacent Motif) - a specific DNA sequence flanking the target.
2) Check if invader DNA sequence is complementary to crRNA.
3) If matched, form R-loop (crRNA binds to complementary DNA sequence) and degrade invader DNA
What are two ways viruses can escape the CRISPR-Cas system?
1) Mutations in viral genetic code to evade immune system.
2) Producing anti-CRISPR proteins that disable CRISPR
What is sgRNA role in the Cas9 system?
An sgRNA (single guide RNA) combines the functions of the crRNA and tracrRNA into a single RNA strand for the Cas9 system
