Advanced Cell Bio
. What does CRISPR stand for?
2. What is the advantage of CRISPR-Cas9?
3. How does CRISPR-Cas9 differ from previous techniques of gene ediƟng?
4. What is the role of gRNA in CRISPR-Cas9?
5. What was the original funcƟon of the CRISPR-Cas system?
6. What known diseases has CRISPR-Cas9 been used for in clinical trials?
7. What is a PAM in the CRISPR-Cas9 system?
8. What are the two enzymes in a Cas protein complex?
9. Once the DNA and RNA are bound, Cas9 is acƟvated to cut the DNA how many bases upstream from the
binding sequence?
10. AŌer Cas9 cuts the target DNA, which repair mechanism creates a mutaƟon and inacƟvates the gene?
11. The CRISPR-Cas9 system in bacteria is akin to our body’s what?
12. The human genome contains approximately 3 billion base pairs and there are about 160 million of the
protospacer adjacent moƟf (PAM) sites in the human genome. Please calculate how oŌen PAM sequences
(the frequency of ‘GG’) present in the mammalian genome in average?
13. What are the advantages of nucleic acid-based vaccines?
14. What is the key advantage of mRNA vaccines over DNA vaccines?
15. What is the drawback of viral vector-based vaccines?
16. Please list an early milestone in the development of mRNA vaccines.
17. What is a key development in the producƟon of mRNA?
18. What is an early advancement in the delivery of nucleic acids into cells?
19. What is the contribuƟon made by Katalin Karikó in advancing the mRNA plaƞorm?
20. What are Drew Weissman's background and experƟse?
21. What is the key finding from the collaboraƟon between Katalin Karikó and Drew Weissman?
22. What is the key finding regarding the acƟvaƟon of dendriƟc cells by in vitro transcribed mRNA?
23. What is the focus of each of the three main companies established by 2010 in the field of mRNA
technology?
24. What is the key aspect of the mRNA-based vaccine development against Middle East Respiratory
Syndrome coronavirus (MERS-CoV)?
25. What is the future directions in mRNA vaccine technology development?
CRISPR stands for "Clustered Regularly Interspaced Short Palindromic Repeats".
The advantage of CRISPR-Cas9 is its efficiency, precision, and versatility in editing genes.
CRISPR-Cas9 differs from previous gene editing techniques by being more precise, easier to use, and less expensive.
gRNA (guide RNA) in CRISPR-Cas9 directs the Cas9 enzyme to the specific target DNA sequence for editing.
The original function of the CRISPR-Cas system in bacteria is as an adaptive immune system to defend against viruses.
CRISPR-Cas9 has been used in clinical trials for diseases such as cancer, sickle cell disease, and beta-thalassemia.
PAM (Protospacer Adjacent Motif) in the CRISPR-Cas9 system is a short DNA sequence that must be present near the target DNA sequence for Cas9 to bind and initiate editing.
The two enzymes in a Cas protein complex are Cas9 (endonuclease) and a nuclease (e.g., Cas1 or Cas3) for processing the DNA.
Cas9 cuts the target DNA three bases upstream from the PAM sequence.
After Cas9 cuts the target DNA, the repair mechanism that creates a mutation and inactivates the gene is non-homologous end joining (NHEJ).
The CRISPR-Cas9 system in bacteria is akin to our body's adaptive immune system.
To calculate the frequency of 'GG' PAM sequences in the mammalian genome, divide the number of PAM sites (160 million) by the human genome size (approximately 3 billion base pairs). The result gives you the average frequency of 'GG' PAM sequences in the mammalian genome.
Advantages of nucleic acid-based vaccines include their safety, rapid development, and potential for broad applicability.
The key advantage of mRNA vaccines over DNA vaccines is their ability to enter the cytoplasm directly without needing to enter the nucleus, reducing the risk of integration into the host genome.
A drawback of viral vector-based vaccines is the potential for pre-existing immunity against the viral vectors, which can reduce their effectiveness.
An early milestone in the development of mRNA vaccines was the demonstration of successful protein expression from exogenous mRNA in cells.
A key development in the production of mRNA is the optimization of RNA synthesis and purification techniques.
An early advancement in the delivery of nucleic acids into cells was the development of lipid nanoparticles (LNPs) for efficient intracellular delivery.
Katalin Karikó made a significant contribution to advancing the mRNA platform by pioneering the use of modified nucleosides in mRNA to reduce its immunogenicity.
Drew Weissman's background and expertise lie in immunology and molecular biology, particularly in the field of mRNA research.
The key finding from the collaboration between Katalin Karikó and Drew Weissman was the discovery that modified nucleosides in mRNA could prevent it from triggering an immune response.
A key finding regarding the activation of dendritic cells by in vitro transcribed mRNA is its ability to induce a strong immune response, making it a promising tool for vaccine development.
The focus of each of the three main companies established by 2010 in the field of mRNA technology varied, but generally included vaccine development, therapeutic applications, and platform technology advancement.
The key aspect of mRNA-based vaccine development against Middle East Respiratory Syndrome coronavirus (MERS-CoV) is its rapid and targeted approach, potentially allowing for faster response to emerging infectious diseases.
Future directions in mRNA vaccine technology development include improving stability, scalability, and delivery methods, as well as expanding applications beyond infectious diseases to include cancer immunotherapy and other therapeutic areas.