CRISPR-Cas9 System

by Georgina Cornwall, PhD

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    00:00 in the plants using this mechanism.

    00:02 So now let us look at a very specific restriction endonuclease system. Recall the restriction endonucleases were used by bacterial cells as the immune system. Well, it turns out that they have some even more specific restriction endonucleases than we had learned about previously. Bacteria will use their endonucleases to cut up viral DNA that has been injected into them to protect themselves from that, but it also turns out that bacteria perhaps have an immune system fairly similar to ours. When we are exposed to a viral pathogen our immune system learns about that has some memory by keeping some of the antigens from those invading particles and next time the invasion happens, we can attack it with more specificity and we are attacking it more quickly to rid ourselves of the invaders much faster. Bacteria turns out to have a very similar mechanism, not only did these particular restriction endonucleases chop up the invading DNA, they also will make a little copy of that invading DNA and store it in their own genome for a later use. How does it work? This is the Cas9 and CRSPR system. CRSPR is an acronym for a really crazy long word. We could have clustered regularly interspaced short palindromic repeats. We understand all for those words separately, but CRSPR is honestly much easier to say and CRSPR is paired with the Cas9 proteins.

    01:50 Let us take an example of a target sequence. We have guide RNA that can now be copied from the bacterial genome that is comparable or pairs with the DNA that was perhaps injected by a virus. Next time that virus comes along, the bacterial cell can produce this guide RNA and it will fit into the Cas9 protein. It works kind of like this. The Cas9 protein has an active site in which this guide RNA that is specifically targeted to the invader will bind. Now the Cas9 protein is on a mission to find any pieces of DNA that match up to this auide RNA sequence. As Cas9 with this target sequence runs around the cell, he is going to bind onto all of the different DNA that are all of the same DNA that much that sequence and hack it up. So a very efficient pair of scissors for cutting DNA into pieces at very specific locations. We can utilize this system in cells. Recently we’ve incorporated these sequences, we have stolen the technology from bacteria once again to now be able to create guide RNA, which allows us, now we have the human genome sequence to actually produce target specific MRNAs, you can actually order them online. There is a whole catalog and you pick your target sequence that mark specific genes are associated with specific genes and actually cut either in the middle of those genes or on the outside of those genes.

    03:43 The CRSPR/Cas9 system specifically targets a DNA sequence could cut in the middle, it can cut wherever we want it to cut because we chose what the target sequence is. This is really a good step for our DNA technologies in being able to introduce sequences. There have been a lot of work going on recently in Parkinson's disease and cystic fibrosis and even in the treatment of HIV attacking the incoming viral DNA. Super exciting field for now. Molecular scissors chop it all up. Now let us move on to how we could amplify

    About the Lecture

    The lecture CRISPR-Cas9 System by Georgina Cornwall, PhD is from the course Biotechnology.

    Included Quiz Questions

    1. …a family of DNA sequences containing pieces of DNA from invading viruses that attack the host bacterium.
    2. …a family of RNA sequences containing pieces of RNA from invading viruses that attack the host bacterium.
    3. …a family of signal protein sequences containing pieces of antigen proteins from invading viruses that attack the host bacterium.
    4. …a family of signal lipoproteins sequences containing pieces of immunogenic proteins from invading viruses that attack the host bacterium.
    5. …a family of signal glycoproteins sequences containing pieces of immunogenic proteins from invading viruses that attack the host bacterium.
    1. CRISPR/Cas 9 system helps in the amplification of viral DNA inside the host cells by providing the adaptive immunity to the incoming virus.
    2. The viral snippets in the CRISPR of a bacterium play a crucial role in bacterial defense system against the bacteriophages.
    3. The pre-designed sequence of the Guide RNA (gRNA) directs the Cas9 nuclease to the right part of the invading DNA molecule for chopping down it.
    4. Cas9 nuclease cleaves a double-stranded DNA at specific sites and leads to activation of DSB repair machinery in the bacterial cells.
    5. CRISPR/Cas9 system can be used as a targeted genome editing tool in the molecular biology protocols.

    Author of lecture CRISPR-Cas9 System

     Georgina Cornwall, PhD

    Georgina Cornwall, PhD

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