All language subtitles for 47031x_MH_Gene_Function_07_Complementation_Test_v1-en

MICHAEL HEMANN: Say you are in the lab

and you've just identified a new strain, right?

And so, we're going to call this strain hisX minus.

So-- and I think this refers to a question that was put

in the chat a little bit earlier on--

so hisX minus is a strain that can't grow

without exogenous histidine.

So it's a histidine auxotroph.

But we don't know anything about what gene

this mutation is in, right?

And you've just developed this in lab,

and you've been working on his3 for a while,

and you want to know--

is hisX minus the same as his3 minus?

Are they simply mutations in the same gene,

or this represent a new gene that's mutated

that causes the same phenotype?

Well how do we figure this out?

Well we figure it out by simply crossing these haploids

together in what we'll call, essentially,

a complementation test.

So we're going to think, essentially,

of two distinct possibilities.

And the first possibility is that hisX minus

is the same as his3 minus.

They're essentially mutations in the same gene.

And so in this case, the diploid genotype

is hisX minus over his3 minus.

Or we can write that, if they're equivalent,

as his3 minus over his3 minus.

And the phenotype of this, of course, is his minus.

So if we're putting the same mutation in opposition

to itself, we're not restoring function.

And so we are now his minus.

Well let's think of, then, the other possibility.

And so the other possibility is hisX minus

is not the same as his3 minus.

And so the resulting diploid genotype

is going to be hisX plus over hisX minus.

And his3 minus over his3 plus.

Why is that?

Because we're dealing here with two distinct genes.

And sort of the inferred phenotype of the haploids

is that you are--

if you are hisX minus, you are his3 plus.

And we can write sort of two genes in series.

So these are two distinct genes-- hisX minus

and his3 plus.

Whereas the other strain would be hisX plus and his3 minus.

So if we're just saying that they're

a mutant for a single gene, we're

assuming they're wild type for all of the others.

And so in each case, you're bringing

a functional copy of his3 and a functional copy of hisX

into this cross.

And so the resulting diploid genotype

is going to be heterozygousity for both of these genes.

And the phenotype is going to be--

yeah.

So there's a question-- are we assuming

there's only one mutation in this synthesis pathway?

We do assume that.

And there are, I think, important ways

that you actually make sure that that's actually the case--

where you actually are maintaining strains

with the idea that there's only sort of single defects.

Now we'll talk about this idea that maybe

there are multiple defects--

I think in a couple of lectures.

But the assumption is, if I say something

is his3 minus, that is the only mutation within this pathway.

OK.

So the resulting, you know, diploid in this case

is going to be his plus.

Because they have one functional copy of hisX,

they have one functional copy of his3,

these are actually compensating for the deficiency

of each other.

And so we're going to introduce this idea of complementation.

And complementation essentially means that the cross between--

or the progeny of a cross between two mutants

is wild type.

Or haploids, or diploids, that have a mutant phenotype

yields a wild type phenotype.

So can you actually restore the function of the gene

deficiency in the other strain?

Can two strains that separately have

a problem, or a phenotypic problem,

restore function in that pathway?

So if you have mutations in this same gene,

you do not complement.

If you have mutations in different genes,

you do complement.

So if you have a study group and you

and your friend know exactly half of the material,

but you know same half of the material,

you're not actually complementing one another,

right?

You're not helping each other out.

You actually have to bring something

that the other person lacks-- in that sense you complement.

So are you actually bringing a functional gene

that complements the gene deficiency in the other.