00:01
All right, so we have three strains.
00:02
A, which is kanamycin resistant, b, which is gentamicin resistant, and a third strain c, which turns purple, but is not resistant.
00:10
So, you want to know which gene, the kanamycin resistance gene or the gentamicin resistance gene, which of these two is on a plasmid.
00:25
So we're trying to solve, right? so, how do we do this? quite simply, we do two matings.
00:35
We do two conjugations experiments.
00:43
Okay, so we conjugate the first one, a and c, which should give us purple, assuming the kanamycin resistance gene is on a plasmid.
00:59
It should give us, if you played out c, it should give us purple kanamycin resistant bacteria.
01:08
This means that c takes up the plasmid that contains this kanamycin resistance gene.
01:26
Same token, we do a second mating of b times c.
01:33
And here, in this case, if gentamicin is plasmid resistant, is the resistance gene on the plasmid, we should get purple gentamicin resistance bacteria, right? so, how do we confirm either of these? both platings, you plate them, put this on kanamycin containing plates.
01:58
I don't know why i'm writing the whole thing.
02:01
Put them on kan plates, see if anything grows.
02:04
This one, put them on gentamicin plates and grow.
02:08
If it grows on here, then we know that the kanamycin resistance gene is on the plasmid.
02:17
If it grows here, then we know that the gentamicin is on the plasmid.
02:22
That's it.
02:25
And the next part of the question is, how would you determine if any of the strains are naturally competent? well, once you figure this out, which is on a plasmid, right? so, let's say, well, it doesn't really matter.
02:46
Figure out which one is on a plasmid.
02:47
Once you've figured out which is on the plasmid, right, whichever one, then whichever one it is, isolate the plasmid.
03:04
Okay? using something called a plasmid prep kit...