00:01
Slide, it talked about the expression of transgene in a host cell produce large amount of protein.
00:10
So what we have is that you have a plasmid.
00:17
The plasmid is considered as a vector because it's going to carry the foreign gene into the host because plasmid is able to go in and out of a host.
00:30
So this plasmid is a dna obviously.
00:33
It's an extra chromosomal dna.
00:37
And on this vector or plasmid, there is a restriction site.
00:42
So you have a lot of enzyme restriction site along this plasmid.
00:48
And one of them is called the bamh1.
00:54
The restriction enzyme cut the dna.
00:58
Now you have the plasmid has the restriction enzyme bamh1.
01:02
And then you also have a foreign dna that also cut with the same enzyme.
01:13
So now you combine them together.
01:16
The plasmid will be cut so that it will be linearized with the bamh1 site on the two ends.
01:26
And then you have the same restriction site bamh1 on a foreign dna.
01:32
I would say the target sequence that you want to put inside of the vector.
01:44
So now you combine the two fragment because they all have the same restriction end, bamh1.
01:53
They'll stick in.
02:02
So this piece go inside.
02:04
So they will stick together.
02:05
This process is called ligation.
02:10
So after ligation, the plasmid will become, will go back into a complete circle.
02:17
But this time, it has a additional piece in it.
02:22
This is the target dna.
02:24
So the bamh1 restriction site is going to go back together by this ligation reaction, which you have enzyme called ligase.
02:33
Ligase is able to put the two fragment together as long as they have the same restriction end.
02:40
They kind of complement each other.
02:43
So they will stick back in.
02:45
So now you have this recombinant plasmid, which you carry this specific target sequence, which is blue.
02:58
Now, once you have the recombinant plasmid, you introduce that with a host e.
03:04
Coli cell.
03:05
So the e.
03:06
Coli has its genomic dna.
03:07
So this is the chromosome dna.
03:15
So what we know about the plasmid is that the plasmid is a extra chromosomal dna that would be able to go inside of e.
03:24
Coli when we have this experiment we call transformation.
03:30
The transformation is an experiment that will focus on the membrane of the e.
03:36
Coli host.
03:38
These are bacteria host.
03:40
Now the plasmid is actually be able to go inside of the e.
03:45
Coli host.
03:46
So as a result, after transformation or successful transformation, that it is able to get into the plasmid.
03:56
This is the recombinant plas host.
04:05
Let's use a different word.
04:07
Let's call transformed.
04:08
It's a better word...