All language subtitles for 57031x_MH_Gene_Function_08_Complementation_Groups_v-en

MICHAEL HEMANN: In this case, because we know nothing about

the gene function itself, right--

we just have a strain that we've isolated that is his-,

we really can't make any assumptions about what it is.

If we actually knew the gene sequence-- knew, for example,

that this his3- actually is just lacking a his3 gene--

then we can make some assumptions

likely about whether it's dominant or recessive,

or what the resulting phenotype of that cross would be.

But if we don't know anything about it

other than it's his deficient, then we actually

have to do the cross.

And I think the important thing here

is that for this extended period of time,

and even now in thinking about this kind of biology

in this kind of strain, you have to have

a way of doing this cross.

So let me give you an example.

So say that you have nine different strains,

and these strains are all his-.

You know nothing about these strains

other than they can't grow in the absence

of exogenous histidine.

What we can do is you can cross all of these together, right?

So you can do pairwise crosses of each of these.

And based on our complementation test, all of the his1-

strains--

and we're just-- that's an arbitrary designation.

That's just the first strain we've identified.

It doesn't place it in the pathway anyways,

but it's the first mutant--

all of the green hist1- strains will complement

all of the other strains.

So they'll complement these black and these red strains.

But they won't complement one another

because you do not complement if you have

mutations in the same gene.

So we can essentially separate these three strains away

from the others and put them in what we'll

call a complementation group.

So the three red strains will be together,

and we can call these his2-.

And these three in black will be put together,

and we'll call them his3-, right?

So they won't complement each other,

but they'll complement all of the others.

And so a complementation group is essentially

a group of mutants that don't complement each other,

suggesting that they are all mutations that

are in the same gene.

And so you can see how this is actually really powerful.

So we know nothing about the gene sequences of his genes.

But by doing this analysis, if we have enough strains,

we can actually identify sets of mutants that have alterations

in the same gene.

And we can even possibly identify

how many enzymes are important in this pathway.

So if we have a ton of mutants, and we

have three complementation groups,

then perhaps and very likely there

are three important enzymes essential

for histidine biosynthesis.

So, again, knowing nothing about the molecular details

and the sequence details of these genes,

we can actually infer a great deal about this biology.

So say we actually had a strain that was dominant, right?

So here, we have a hisD strain, right?

How many strains will this hisD strain

complement if it is a dominant strain that

doesn't produce histidine?

Most of you say zero, and that's, in fact, correct.

So in this strain, or in this cross,

if we're actually crossing a dominant strain

to all of these recessive strains,

then the heterozygote, because it's dominant,

is going to show the his- phenotype.

So if you do this kind of complementation test

with a dominant strain, then it will complement nothing, right?

And, in fact, you can't infer anything about this gene other

than it is a dominant allele.

It could be a dominant allele of his1, 2, or 3.

It could be an allele of something else.

So you can't place it into a complementation

group with anything else.

But you can infer a relationship between these.

And so, for the recessive strains,

we can actually cluster them into a complementation group.

And this has actually been a really effective strategy

in a lot of human conditions.

So if you have, for example, a peroxisomal biogenesis

disorder, you can't make peroxisomes.

There are 15 different kinds of mutations

in different genes all involved in peroxisomal biogenesis.

And by actually fusing cells together

from people with these conditions,

you can do the same test.

It's essentially a diploid-cell way

of doing a cross, where you fuse two cells together,

and you see, does it actually make peroxisomes?

And you can separate these into complementation groups

that correspond to each gene.

And that becomes really important if you're

going to do mapping studies, like we'll talk about,

because you want to know, am I mapping to a specific gene?

So if you're looking at a bunch of different genes,

then you're going to get a bunch of different locations

in the genome.

If you know that these are all mutations in the same gene,

then you can focus your study on these

and identify a locus much more effectively.