Download Complementation Help - Biological Sciences

BIOLOGY 207 - Dr. McDermid
Complementation Help
Complementation is a fundamental principle in genetics and its understanding is needed to appreciate many
aspects of genetics.
Definitions: (Modified from "A Dictionary of Genetics" 4th Edition by RC King & WD Stansfield, 1990.)
Complementation - the appearance of a wild type (normal) phenotype in a diploid organism or cell
containing two different mutations. One can conclude that the mutations are in different genes (non-allelic)
because the phenotype is wild type.
Non-complementation or lack of complementation - the appearance of a mutant
phenotype in an diploid organism or cell containing two different mutations. One can conclude that the mutations
are in different alleles of the same gene (allelic) because the phenotype is mutant.
Nomenclature - Prior to a complementation test we will be unsure of the relationship between the two
mutations (allelic or non-allelic), so the nomenclature needs to be ambiguous. Therefore, "m1" and "m2" are
used and not "a" and "b", which implies two loci, or a1 and a2, which implies alleles.
Complementation test - the introduction of two mutations into the same cell to see if the mutations in
question (e.g. "m1" and "m2") occurred in the same gene or in different genes.
If the mutations are allelic then a mutant phenotype will result and the genotype of the hybrid may be
symbolized as !m1/m2!.
If the mutations are nonallelic then the phenotype will be wild type and the genotype of the hybrid may be
symbolized as !+/ m1 ; +/ m2.
If the wild type phenotype is expressed, each normal allele "makes up for" or “rescues” or "complements" the
defect in the corresponding mutant allele.
Complementation test:
In Haploid species (e.g. yeast)
In Diploid species (e.g. Drosophila,
mice, plants, etc.)
Introduction:
You begin with two (or more) independently derived
mutations in strains of haploid yeast that have similar mutant
phenotypes. In this example we will call the two strains "m1"
and "m2", for mutation#1 and mutation#2. The mutant
phenotype could be colony colour, auxotrophy of the same
compound, or any other character that distiguishes it from
wild type.
The question to be answered in a complementation test
is:
Are these two (or more) strains mutant in the same gene or
are they mutant in different genes? That is are m1 and m2
allelic or non-allelic mutations?
Introduction:
You begin with two (or more) independently derived
mutations in strains of diploid speices that have similar
mutant phenotypes. In this example we will call the two
strains "m1" and "m2", for mutation#1 and mutation#2. The
phenotype could be eye colour or any other character that
distinguishes it from wild type.
The question to be answered in a complementation
test is:
Are these two (or more) strains mutant in the same gene
or are they mutant in different genes? That is are m1 and
m2 allelic or non-allelic mutations?
TEST:
TEST:
We can "cross" yeast strains of differing mating types, a and We "cross" males of one strain (one mutation) with female
a (alpha). (This is similar to males and female in diploid
of another strain (a different mutation). The resulting
species.) The cross is done by putting the two strains, with
progeny are tested or examined for their phenotype the different mutations (and different mating types) together mutant or wild type?
on a petri dish containing mating medium.
When mating occurs the two haploid cells fuse to form one
diploid cell containing both mutations. We can now
propagate
diploid cell to make a diploid
cell strain
that
Biol207 this
Dr. McDermid
Lecture#3
-supplement
page 1
we can test or examine for its phenotype - mutant or wild
type?
diploid cell containing both mutations. We can now
propagate this diploid cell to make a diploid cell strain that
we can test or examine for its phenotype - mutant or wild
type?
One of two outcomes is possible from a cross of
Strain#1 (x) Strain#2
Both are mutant in the
same gene.
Allelic mutations
Strain#1 (x) Strain#2
m1 (x) m2
Mutations are in different
genes.
Non-allelic mutations
Strain#1 (x) Strain#2
m1 ;+!(x) !+!;m2
Fuse cells to make
diploid:
m1/m2
Fuse cells to make diploid:
m1!/+ ; +!/ m2
The phenotype is:
The phenotype is:
MUTANT!
Defect in one
strain is NOT
complemented by
the other.
They are two
mutant alleles in
the same gene
locus.
NONcomplementation!
One of two outcomes is possible from a cross of
Strain#1 (x) Strain#2
Both are mutant in the
same gene.
Allelic mutations
Mutations are in different
genes.
Non-allelic mutations
Strain#1 (x) Strain#2
m1/m1 (x) !m2/m2
Each diploid parent
produces haploid
gametes:
m1!!!and !! m2
These fuse to make
Diploid progeny:
m1/m2
Strain#1 (x) Strain#2
m1/m1;+/+ (x)!!+/+;m2/m2 !
Each diploid parent
produces haploid gametes:
m1!;+ !!!and !!!+ ;m2
These fuse to make
Diploid progeny:
m1!/+ ; +!/ m2
The phenotype is:
MUTANT!
WILD TYPE!
Defect in one strain Defect in one
is complemented by strain is NOT
the other.
complemented by
the other.
They are mutants in
They are two
two different gene mutant alleles in
loci, one in m1 the
the same gene
other in m2.
locus.
NONComplementation!
complementation!
The phenotype is:
WILD TYPE!
Defect in one strain
is complemented
by the other.
They are mutants in
two different gene
loci, one in m1 the
other in m2.
Complementation!
Conclusion:
The outcome of the complementation test (wild type or mutant) permits you to distinguish between allelic vs. nonallelic mutations.
If the two mutations were allelic then you might change the mutants' symbol to reflect that. For example they might
become m1 and m2, with the superscripts designating different mutant alleles.
If the two mutations were non-allelic then you might change the mutants' symbols as well. For example if the
mutations were auxotrophic for arginine then you might call them the arg1 locus and the arg2 locus, with the
mutant alleles designated as a "minus" symbol : !!!arg1- !!and !!!arg2- .
Make sure you understand the symbols' use and meaning here.
Note:
Note:
Since the initial mutant cells used here are haploid there
Only recessive mutations can be tested for
is no "dominance or recessiveness".
complementation since dominant mutations would
Only two strains can be tested at once but through a
show a mutant phenotype in all the progeny with
series of tests the relationship among a series of
the dominant mutant allele, independent of
strains can be determined.
whether or not there is one or two genes involved.
Independently derived mutant strains are those that are
Only two strains can be tested at once but through a
originally mutated at a different time or place and so
series of tests the relationship among a series of
could not be due to the same mutational event.
strains can be determined.
Independently derived mutant strains are those that are
originally mutated at a different time or place and
so could not be due to the same mutational event.
Biol207 Dr. McDermid
Lecture#3 -supplement
page 2
Questions to test your understanding
1. The following results were obtained from a series of
complementation crosses for auxotrophic yeast mutants a - f.
a b c d e f
a
- + + - + +
b
+ - + + - c
+ + - + + d
- + + - + +
e
+ - + + - f
+ - - + - - = no growth + = growth
What can be said about each of these mutations and the
gene(s) in which they reside? Explain.
2. Four pure-breeding, mutant, diploid, plant lines (a,
b, c, and d) were developed that had white pedals in a
plant which normally has purple pedals (wild type = +).
These four lines, when crossed among themselves
(complementation tests) and to wild type, gave the
following results:
+
a
b
c
d
+ purple purple white
purple
purple
a purple white white
purple
white
b white white white
white
white
c purple purple white
white
purple
d purple white white
purple
white
a. What is the "dominance/recessive" relationship of
each of a, b, c, and d relative to wild type?
!
b. Explain the relationships (complementation groups)
amongst the five alleles in the four mutant lines and wild
type.
c. How many genes are involved here.
!
Answer:
There are three genes:
Mutations a and d fail to complement and therefore
represent alleles of one gene.
Mutations b and e fail to complement and therefore
represent alleles of one gene.
Mutations a/d complement b/e and therefore represent
different genes.
Mutation c complements a/d and b/e and therefore
represents a third gene.
The strain carrying mutation f must actually have two
mutations, one in the "b/e" gene and one in the "c"
gene, or be a deletion including both genes.
Answers:
a) b is dominant and a, c, and d are recessive.
b) c complements a and d
a and d fail to complement
b is unknown because it is a dominant mutation
c) At least two (a/d, c). Because b is dominant we
can't tell if it represents a different gene or one of
the two known genes.
Lecture notes: Copyright © 1996-2002! Locke/McDermid and the Department of Biological Sciences, University of Alberta,
Edmonton, Alberta, Canada.
Biol207 Dr. McDermid
Lecture#3 -supplement
page 3