Genetics Lecture Notes

7.03 2005

Lectures 1 – 2
 Lecture 1

We will begin this course with the question: What is a gene?

This question will take us four lectures to answer because there are actually several
different definitions that are appropriate in different contexts.

We will start with a physical definition of the gene. Conceptually this is the simplest
and it will give me an excuse to briefly review some of the molecular biology that you
probably already know.

Genes are made of DNA

For this course we will mostly think of DNA as an information molecule rather than
a chemical substance.

In 1953, Watson and Crick deduced that the structure of DNA was a double helix. It
was not the helical structure per se, but the discovery of complementary base pairing
that revealed how information could be encoded in a molecule and how this information
could be exactly duplicated each cell division.

Replication.
In order to extract information from the DNA, the cell again uses the complementary
base-pairing to make a copy of the information copied onto an RNA molecule. This is
known as Transcription. RNA is chemically less stable than DNA and mRNA can be
thought of as a temporary copy of DNA’s information.

Transcription

Translation

Folded Proteins:
enzymes
structural proteins
membrane channels
hormones, signaling molecules
Gene: DNA segment needed to make a protein

Genes are typically 103 - 104 base pairs in size although they can be much larger. For
example, the human dystrophin gene is 2 x 106 base pairs.

E. coli has about 4,200 genes which isn’t very many considering that at least 1,000
different enzymes are needed to carry out just the basic biochemical reactions in a
cell. The smallest genome for a free-living organism (i.e. a cell, not a virus) is that of
the bacterium Mycoplasma genetalium which encodes only 467 genes. Humans are at
the other end of the spectrum of complexity and have about 20,000 - 25,000 genes.
In the demonstration in class you see how a mutation in the Shibire gene in the fly
Drosophila gives a heat sensitive protein that is required for synaptic transmission.
When the flies that carry this mutation are warmed by the projector lamp they
become paralyzed.

Gene — Protein — Cellular function — Organismal function

(Shibire) (Dynamin) (Vesicle recycling) (Neuromotor activity)

This example illustrates two powerful aspects of genetic analysis. First, we can follow
microscopic changes in the DNA such as the temperature sensitive mutation in the
Shibire gene as they are revealed by the macroscopic consequences of the mutation
such as a paralyzed fly. Second, we have a very precise way of studying the function
of individual proteins by examining the consequences of eliminating just that one
protein function in an otherwise normal organism.

Alleles: different versions of the same gene

Often alleles are referred to as mutants but actually this usage is often incorrect
particularly when we discuss naturally occurring variants in a population.

Mutation: an altered version of a gene when we have “witnessed” the alteration but
not when it is preexisting in the population.

Genotype: all alleles of an individual

Wild-type: defined standard genotype

The concept of wild-type is used as a defined reference for organisms where we can
do breeding experiments. Of course there is no realistic way to define a standard
genotype for humans, therefore “wild type” has no meaning when we discuss human
genetics.

The physical definition of the gene is a very good one but there are many instances
where we wish to study genes whose DNA sequences are not known. For example, say
we have isolated a new mutant fly that is also paralyzed and we want to know whether
this mutation is also in the Shibire gene. We will see in the next several lectures that
we can answer this question without knowledge of the DNA sequence either by a test
for gene function known as a complementation test or by a test of the chromosomal
position of the mutation by recombinational mapping. In practice, these other ways of
defining genes by function or by position are often much more useful than a definition
based on the DNA sequence.
 Lecture 2

In this lecture we are going to consider experiments on yeast, a very useful organism for
genetic study. Yeast is more properly known as Saccharomyces cerevisiae, which is the
single-celled microbe used to make bread and beer. Yeast can exist as haploids of either
mating type a (MATa) or mating type a (MATa). Haploid cells of different mating type when
mixed together will mate to make a diploid cell.

Haploids and diploids are isomorphic – meaning that a given mutation will cause essentially the
same change in haploid and diploid cells. This allows us to look at the effect of having two
different alleles in the same (diploid) cell.

All yeast needs to grow are salts, minerals, and glucose (minimal medium). From these
compounds, yeast cells can synthesize all of the molecules such as amino acids and
nucleotides that are needed to construct a cell. The synthesis of complicated molecules
requires many enzymatic steps. When combined, these enzymatic reactions constitute a
biochemical pathway

Consider the pathway for the synthesis of the amino acid histidine.

A B C D histidine Protein

Enzyme: 1 2 3 4

Each intermediate compound in the pathway is converted to the next by an enzyme. For
example, if there is a mutation in the gene for enzyme 3 then intermediate C can not be
converted to D and the cell can not make histidine. Such a mutant will only grow if histidine
is provided in the growth medium.
This type of mutation is known as an auxotrophic mutation and is very useful for
genetic analysis.
growth on minimal growth on minimal + histidine

His+ (wild-type) + +
His– – +
Phenotype: All traits of an organism (with an emphasis on trait under investigation)

Homozygote: diploid with two like alleles of same gene

Heterozygote: diploid with two different alleles of same gene

Recessive Allele: trait not expressed in heterozygote

genotype phenotype Mate to : diploid genotype diploid phenotype

MATa His3– His– MATa His3– His3–/His3– His–

MATa His3– His– MATa His3+ His3–/His3+ His+

Based on the His– phenotype of the His3–/His3+ heterozygote, we deduce that His3–
is recessive to wild type.

Let’s consider a different kind of mutation giving resistance to copper that occurs in
a gene known as CUP1.

genotype phenotype Mate to : diploid genotype diploid phenotype

MATa Cup1r copper resistant MATa Cup1+ Cup1r/Cup1+ copper resistant

Dominant Allele: trait is expressed in heterozygote

Cup1r is dominant to wild-type (Cup1+).

The terms dominant and recessive are simply shorthand expressions for the results
of particular experiments. If someone says a particular allele is dominant that means
that at some point they constructed a heterozygous diploid and found that the trait
was expressed in that diploid.
Note: Sometimes an allele will have more than one phenotype and may be recessive for one
and dominant for another. In such cases, the phenotype must be specified when one is
making statements about whether the allele is dominant or recessive. Consider for
example, the allele for sickle cell hemoglobin in humans designated Hbs. Heterozygous
individuals (Hbs/Hba) are more resistant to malaria, thus Hbs is dominant for the trait of
malaria resistance. On the other hand, Hbs/Hba heterozygotes do not have the
debilitating sickle cell disease, but Hbs/Hbs homozygous individuals do. Therefore, Hbs is
recessive for the trait of sickle cell disease.

Once we find out whether an allele is dominant or recessive, we can already infer important
information about the nature of the allele. The following conclusions will usually be true.

Recessive alleles usually cause the loss of something that is made in wild type

Dominant alleles usually cause increased activity or new activity

It turns out that the Cupr allele actually carries more copies of the gene for a copper
binding protein and therefore increases the activity of the gene.

Last lecture we defined the gene structurally as the DNA needed to encode a protein. We
can now define a gene in a new way based on its function. Using the phenotypic difference
between wild type and a recessive allele we can use a Complementation test to determine
whether two different recessive alleles are in the same gene.

Say you isolate a new recessive histidine requiring mutation that we will call HisX–. In
principle, this mutation could be in His3 or it could be in any of the other genes in the
histidine biosynthetic pathway. In order to distinguish these possibilities we need a test
to determine whether HisX is the same as His3.

To carry out a complementation test, one simply constructs a diploid carrying both the
His3– and HisX– alleles.

An easy way to do this would be to mate a MATa HisX– strain to a MATa His3– strain.

possibility genotype of diploid phenotype of diploid complementation

HisX = His3 His3–/His3– His– No

HisX π His3 His3–/His3+, HisX–/HisX+ His+ Yes

This test only works for recessive mutations. Think about what the outcome would be if
HisX– were dominant.

The complementation test can be thought of in the following way. If I have an allele with
an observable phenotype whose function can be provided by a wild type genotype (i.e., the
allele is recessive) — I can ask whether the function that was lost because of the
recessive allele can be provided by another mutant genotype. If not, the two alleles must
be defective in the same gene. The beauty of this test is that the trait can serve as a
read-out of gene function even without knowledge of what the gene is doing at a
molecular level.
Gene: The fundamental unit of heredity which can be defined in three ways. i) A gene can be
defined in molecular terms as a segment of DNA carrying the information necessary to express a
complete protein or RNA molecule including the promoter and coding sequence. ii) A gene can be
defined by function with a group of recessive mutations that do not complement each other. iii) A
gene can be defined by position with a single-locus segregation pattern in a cross between lines with
different alleles. Examples are a 1:3 phenotypic ratio in the F2 generation in a cross between
diploid organisms or a 2:2 segregation pattern in yeast tetrad analysis.

Alleles: Different versions of the same gene.

Locus: The site on a chromosome where a gene is located. Usually defined by recombinational
mapping relative to neighboring loci.

Genotype: The allelic constitution of an individual, usually with emphasis on the gene or genes under
examination.

Phenotype: All of the traits or characteristics of an organism, usually with emphasis on traits
controlled by the gene or genes under examination.

Wild-type: A standard genotype that is used as a reference in breeding experiments. Note that
for human crosses there is no standard genotype and the concept of wild-type is therefore not
meaningful.

Haploid: A cell or organism with one set of chromosomes (1n).

Diploid: A cell or organism with two sets of chromosomes (2n).

Homozygous: The condition of having two like alleles in a diploid.

Heterozygous: The condition of having two different alleles in a diploid.

Dominant allele: An allele that expresses its phenotypic effect or trait in the heterozygous state.

Recessive allele: An allele whose phenotypic effect or trait is not expressed in a heterozygous
state.

Incomplete dominance: The case where a heterozygote expresses a phenotype intermediate

Complementation test: A test of gene function where two genotypes with recessive alleles are
combined by a cross to test whether the genotype of one parent can supply the function absent in
the genotype of the other parent.

F1: First generation produced by interbreeding of two lines.

F2: Generation produced by interbreeding of F1 individuals.

Incomplete penetrance: Cases where certain alleles are not always expressed to give observable
traits because of other environmental or genetic influences.

True-breeding: Refers to a line of individuals that on intercrossing always produce individuals of