In this video, we will be talking about CRISPR
and RNAi.
We will be going over some background for
the video, the basics of each method of gene
therapy, how they do their job, how we choose
between them, and which one I would choose.
First let's get into background.
As you can see I have some information on
this slide and I will be touching on each
of the bullet points.
We will be going over two methods of gene
therapy, RNAi and CRISPR.
RNAi stands for RNA interference and CRISPR
starts for cluster regulatory interspaced
short palindromic repeats.
In the hypothetical situation that we will
be going over in this presentation, we will
be trying to eliminate the production for
a protein produced by, what is called, gene
A. To understand which of these methods would
be most effective, and which one I would choose,
let's look at the basics for each method.
For the basics, let's start with CRISPR.
CRISPR is a naturally occuring system that
was used to protect cells from viruses.
The CRISPR mechanism is composed of a Cas
9 protein, which cuts DNA, and guide RNA,
which aligns the mechanisms.
Now, for RNAi, it is important to realize
this is a mechanism and not a type of RNA.
The two main types of RNA in RNAi are siRNA,
which is small-interference RNA, and microRNA.
This censors gene expression in the cell,
and could potentially shut off the gene function.
With the basics out of the way, let’s go
into more depth about how these mechanisms
work.
Again, let's start with CRISPR, which is composed
of guide RNA and a Cas 9 protein.
The CRISPR mechanism has a specific guide
RNA which then locks onto a complementary
DNA sequence.
The Cas 9 protein then cuts the DNA.
The cell then repairs the DNA and allows the
gene function to change and inhibits any further
protein production.
RNAi is also naturally occuring because the
siRNA and the microRNA are naturally occurring
in the body.
When a double stranded RNA is produced, an
enzyme called Dicer cuts the RNA, which is
then picked up by RISC, RNA induced silencing
complex.
The RISC then finds matching mRNA that complements
the picked up RNA.
The mRNA is then cut and the cell degrades
it.
This removes the message the mRNA was carrying
preventing protein production from the gene.
With the function explained, let's look at
how to choose.
Choosing between these two depends on what
you want the outcome to be.
CRISPR does gene knockout, which is when the
gene you are trying to inhibit is knocked
out of the genome.
The CRISPR mechanism has less off-target effects,
which makes it fairly easy to use.
Finally, CRISPR targets DNA.
However, in RNAi, mRNA is targeted.
Along with this difference RNAi s the effect
of gene knockdown which limits the effect
of a gene, but rarely does it eliminate it
completely.
Also with RNAi off-target effects are more
likely to occur because the complementation
of the RNA isn’t strict, and it can have
this effect in other mRNA.
Between these two methods, I would choose
CRISPR because it is easy to use and has less
off-target effects which allows for more precise
cutting because of the Cas 9 protein, which
completely inhibits gene function.
I has also been constantly improved and has
been used more within this field, thank you.
