Applications of the CRISPR-Cas9 system

Applications of the CRISPR-Cas9 system

Nowadays, there are many applications that apply the CRISPR-Cas9 tools, including gene knockout, knockin SNPs, insertions and deletions in cell lines and animals. If you interest, visiting the website http://www.origene.com/CRISPR-CAS9/. Here, they are some applications of CRISPR/Cas9 for therapeutics,

CRISPR/Cas9 for promising treatment of HIV infection

Currently, there are many researches study about how to use the CRISPR/Cas9 as a tool to fight HIV that infects the cell.

Learn more in article: 

-HIV overcomes CRISPR gene-editing attack
-CRISPR/Cas9-Derived Mutations Both Inhibit HIV-1 Replication and Accelerate Viral Escape
-In Vivo Excision of HIV-1 Provirus by saCas9 and Multiplex Single-Guide RNAs in Animal Models

CRISPR/Cas for promising treatment of cancer

CRISPR/Cas could be used as the tool to study the cancer for therapy.

Learn more in article:

-Targeting genomic rearrangements in tumor cells through Cas9-mediated insertion of a suicide gene

CRISPR/Cas for development the antimicrobials

CRISPR/Cas could be used as the tool to kill the specific bacteria that are harmful but not affect the harmless bacteria. The application bases on the process of bacteria that generate the short RNA sequence to guide the Cas9 to fight the bacteriophage by removing its DNA [1]. Here, they are some articles that employ the CRISPR/Cas tool in study of antimicrobials, 

Learn more in article: 

-Exploiting CRISPR-Cas nucleases to produce sequence-specific antimicrobials

-Sequence-specific antimicrobials using efficiently delivered RNA-guided nucleases 


References

1. https://www.nature.com/news/antibiotic-alternatives-rev-up-bacterial-arms-race-1.17621

2. The page of Biology Beyond Nature: ชีววิทยาเหนือธรรมชาติ on facebook

CRISPR/Cas9 videos

Let’ watch the CRISPR/Cas9 videos

• https://youtu.be/2pp17E4E-O8
• https://www.youtube.com/watch?v=0dRT7slyGhs&index=1&list=PL4_fJegcjcJ_doqefkW1hoIee_sgivMr2
• https://www.youtube.com/watch?v=SuAxDVBt7kQ&feature=youtu.be
• https://www.youtube.com/watch?v=MnYppmstxIs
• https://www.youtube.com/watch?v=sweN8d4_MUg
• https://www.youtube.com/watch?v=9_v0dqquUfY
• https://www.youtube.com/watch?v=O3e2_Ctty_M
• https://www.youtube.com/watch?v=UfA_jAKV29g
• https://www.youtube.com/watch?v=sp9eCYHcP5I&t=5s
• https://www.youtube.com/watch?v=iWL_hYM1sq4

Conclusions

CRISPR-Cas9 is the tool for editing the genome at desired target in many organisms. CRISPR-Cas system is firstly discovered in bacteria and archaea as the adaptive immune systems to destroy invading viruses or bacteriophage by recording and targeting their DNA sequences. This system requires a guide RNA to guide the endonucleases (Cas) to cleave foreign genetic elements in particular genome sequence. At present this technology can be applied in numerous applications from prokaryotes to mammals. In the future, this technology will be used to support and resolve the hard disease in various organisms, especially, in human.

The CRISPR-Cas systems

The CRISPR-Cas systems 

The CRISPR-Cas systems are subdivided into Class 1 (Types I, III and IV) and Class 2 (Types II, V, VI) [1]. Each system comprises of specific endonuclease proteins (Cas) and a guide RNA molecule [2]. The guide RNA molecule is responsible to guide the Cas protein to cut and remove foreign DNA in a specific location [3].


The type II CRISPR/Cas9 system (Figure 4), are the simplest one among the three types and original for the genome editing technology. Its processes involved, firstly, the CRISPR array contains repeat regions separated by spacer regions derived from phage or invading pathogens. These are transcribed into a precursor (pre-crRNA). After that, the repeats hybridize with anti-repeat sequences within the tracrRNA. Then, the guide RNA molecule binds to a recombinant form of Cas9 protein that has DNA endonuclease activity (RNaseIII) and the resulting complex will cause target-specific double-stranded DNA cleavage [2,4].

Figure 4. The process of type II CRISPR/Cas9 system [2].

References

1. Makarova, K.S., Wolf, Y.I., Alkhnbashi, O.S., Costa, F., Shah, S.A., Saunders, S.J., Barrangou, R., Brouns, S.J.J., Charpentier, E., Haft, D.H., Horvath, P., Moineau, S., Mojica, F.J.M., Terns, R.M., Terns, M.P., White, M.F., Yakunin, A.F., Garrett, R.A., van der Oost, J., Backofen, R., Koonin, E.V., 2015. An updated evolutionary classification of CRISPR-Cas systems. Nat Rev Micro 13, 722-736.

2. Oude Blenke, E., Evers, M.J., Mastrobattista, E., van der Oost, J., 2016. CRISPR-Cas9 gene editing: Delivery aspects and therapeutic potential. J Control Release 244, 139-148.

3. https://www.broadinstitute.org/what-broad/areas-focus/project-spotlight/questions-and-answers-about-crispr

4. http://www.clontech.com/US/Products/Genome_Editing/CRISPR_Cas9/Resources/About_CRISPR_Cas9

CRISPR/Cas9

What is CRISPR/Cas9

CRISPR stands for clustered regularly interspaced short palindromic repeats, and Cas stands for CRISPR- associated proteins. CRISPR has been firstly discovered in archaea and after that in bacteria by Francisco Mojica (Figure 1), a scientist at the University of Alicante in Spain [4]. Consequently, both scientists women, they are Jennifer Doudna and Emmanuelle Charpentier discovered the CRISPR-Cas9 technology for genome editing. In 2011, Feng Zhang optimized the CRISPR system to work in human cell, (Figure 2).

Figure 1. Francisco Mojica [6]

Figure 2. Jennifer Doudna, Emmanuelle Charpentier, and Feng Zhang [5] 

The CRISPR is the repeating sequences on the DNA sequences and will be disrupted by spacer sequences from virus attack. Cas protein is the enzyme to cut the DNA sequences, particularly, Cas9 is one of the enzymes produced by the CRISPR system [3]. Both of the CRISPR and Cas9 refer to the CRISPR/Cas9 system. This system is a part of adaptive immune system of bacteria and archaea, (Figure 3) to defend against the invading viruses or bacteriophage by recording and targeting their DNA sequences [2,4].

Figure 3. Adaptive immune system of bacteria and archaea [1].

References

1. Gibson, G.J., Yang, M., 2017. What rheumatologists need to know about CRISPR/Cas9. Nat Rev Rheumatol 13, 205-216.

2. Mojica, F.J., Montoliu, L., 2016. On the Origin of CRISPR-Cas Technology: From Prokaryotes to Mammals. Trends Microbiol 24, 811-820.

3. Ran, F.A., Hsu, P.D., Wright, J., Agarwala, V., Scott, D.A., Zhang, F., 2013. Genome engineering using the CRISPR-Cas9 system. Nat. Protocols 8, 2281-2308.

4. https://www.broadinstitute.org/what-broad/areas-focus/project-spotlight/questions-and-answers-about-crispr

5. http://labiotech.eu/wp-content/uploads/2016/09/2792938304_aac596be17_o.jpg

6. http://spainculturescience.co.uk/wp-content/uploads/2017/03/CRISPR-ScotlandAbril20172-Eventos-SRUKCERU-Boletin-OCSA.jpg