Transcript Chapter 17

17
Genomes
Figure 17.7 Synthetic Cells
17 Genomes
17.1 How Are Genomes Sequenced?
17.2 What Have We Learned from
Sequencing Prokaryotic Genomes?
17.3 What Have We Learned from
Sequencing Eukaryotic Genomes?
17.4 What Are the Characteristics of the
Human Genome?
17.5 What Do the New Disciplines of
Proteomics and Metabolomics
Reveal?
17 Genomes
No other mammal shows as much
phenotypic variation as dogs. The Dog
Genome Project sequences entire
genomes of different breeds and
identifies genes that control specific
traits, such as size.
Opening Question: What does dog
genome sequencing reveal about other
animals?
17.1 How Are Genomes Sequenced?
Genome sequencing: determine the
nucleotide base sequence of an entire
genome.
The information is used to:
• Compare genomes of different
species to trace evolutionary
relationships
• Compare individuals of the same
species to identify mutations that
affect phenotypes
17.1 How Are Genomes Sequenced?
• Identify genes for particular traits,
such as genes associated with
diseases
The Human Genome Project was
proposed in 1986 to determine the
normal sequence of all human DNA.
Methods used were first developed to
sequence prokaryotes and simple
eukaryotes.
17.1 How Are Genomes Sequenced?
To sequence an entire genome, the
DNA is first cut into millions of small,
overlapping fragments.
Then many sequencing reactions are
performed simultaneously.
17.1 How Are Genomes Sequenced?
High-throughput sequencing uses
miniaturization techniques, principles
of DNA replication, and polymerase
chain reaction (PCR).
It is fully automated, rapid, and
inexpensive.
Figure 17.1 DNA Sequencing
17.1 How Are Genomes Sequenced?
1. DNA is cut into small fragments
physically or using enzymes.
2. The fragments are denatured using
heat, separating the strands.
3. Short, synthetic oligonucleotides are
attached to each end of each
fragment, and these are attached to
a solid support.
17.1 How Are Genomes Sequenced?
4. Fragments are amplified by PCR.
Sequencing:
1. Universal primers, DNA polymerase,
and the 4 nucleotides (dNTPs,
tagged with fluorescent dyes) are
added.
2. One nucleotide is added to the new
DNA strand in each cycle, and the
unincorporated dNTPs are removed.
17.1 How Are Genomes Sequenced?
3. Fluorescence color of the new
nucleotide at each location is
detected with a camera.
4. Fluorescent tag is removed and the
cycle repeats.
17.1 How Are Genomes Sequenced?
Then the sequences must be put
together.
The DNA sequence fragments, called
“reads,” are overlapping, so they can
be aligned.
17.1 How Are Genomes Sequenced?
Example: Using a 10 bp fragment, cut
three different ways:
TG, ATG, and CCTAC
AT, GCC, and TACTG
CTG, CTA, and ATGC
The correct order is ATGCCTACTG.
Figure 17.2 Arranging DNA Fragments
17.1 How Are Genomes Sequenced?
The field of bioinformatics was
developed to analyze DNA sequences
using complex mathematics and
computer programs.
Figure 17.3 The Genomic Book of Life
17.1 How Are Genomes Sequenced?
Genome sequence information is used
in two research fields:
• Functional genomics—sequence
information is used to identify
functions of various parts of
genomes:

Open reading frames—gene coding
regions
17.1 How Are Genomes Sequenced?

Amino acid sequences, deduced
from sequences of open reading
frames

Regulatory sequences, such as
promoters and terminators.

RNA genes

Other noncoding sequences
17.1 How Are Genomes Sequenced?
• Comparative genomics:
comparison of a newly sequenced
genome with sequences from other
organisms.
This provides more information about
functions of sequences and can be
used to trace evolutionary
relationships.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
The first life forms to be sequenced
were the simplest viruses with small
genomes.
The first complete genome sequence of
a free-living cellular organism was for
the bacterium Haemophilus influenzae
in 1995.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Bacterial and archaeal genomes are:
• Small, and usually organized into a
single chromosome
• Compact—85% is coding sequences
• Usually do not have introns
• Have plasmids, which may be
transferred between cells
Table 17.1
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Functional genomics:
• H. influenzae chromosome has
1,727 open reading frames.
• When it was first sequenced, only
58% coded for proteins with known
functions.
• Since then, the roles of many other
proteins have been identified.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
• Highly infective strains of H.
influenzae have genes for surface
proteins that attach the bacterium to
the human respiratory tract.
• These surface proteins are now a
focus of research on treatments for
H. influenzae infections.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Comparative genomics:
• M. genitalium lacks enzymes to
synthesize amino acids, so it must
obtain them from the environment.
• E. coli has 55 genes that encode
transcriptional activators, whereas
M. genitalium has only 7—a relative
lack of control over gene expression.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Genome sequencing provides insights
into microorganisms that are
important in agriculture and medicine.
Surprising relationships between
organisms suggests that genes may
be transferred between different
species.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Rhizobium bacteria form symbiotic
relationships with plants. The bacteria
fix N into forms useable by plants.
Sequencing has identified genes
involved in successful symbiosis, and
may broaden the range of plants that
can form these relationships.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
E. coli strain O157:H7 causes illness in
humans.
1,387 genes are different from those in
the harmless strains of this bacterium,
but are also present in other
pathogenic bacteria, such as
Salmonella.
This suggests genetic exchange
among species.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Severe acute respiratory syndrome
(SARS) was first detected in southern
China in 2002 and rapidly spread in
2003.
Isolation and sequencing of the virus
revealed novel proteins that are
possible targets for antiviral drugs or
vaccines.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Genome sequencing of organisms
involved in global ecological cycles:
• Some bacteria produce methane, a
greenhouse gas, in cow stomachs.
• Others remove methane from the air.
Understanding the genes involved in
methane production and consumption
may help us slow the progress of
global warming.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Traditionally, microorganisms have
been identified by culturing them in
the laboratory.
Now, PCR and DNA analysis allow
microbes to be studied without
culturing.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
DNA can also be analyzed directly from
environmental samples.
Metagenomics—genetic diversity is
explored without isolating intact
microorganisms.
Sequencing is used to detect presence
of known microbes and previously
unidentified organisms.
Figure 17.4 Metagenomics
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
It is estimated that 90% of the microbial
world has been “invisible” to biologists
and is only now being revealed by
metagenomics.
The increased knowledge of the
microbial world will improve our
understanding of ecological processes
and better ways to manage
environmental problems.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Transposable elements
(transposons) are DNA segments
that can move from place to place in
the genome or to a plasmid.
If a transposable element is inserted
into the middle of a gene, it will be
transcribed, and result in abnormal
proteins.
Figure 17.5 DNA Sequences That Move (A)
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Composite transposons:
transposable elements located near
one another will transpose together
and carry the intervening DNA
sequence with them.
Genes for antibiotic resistance can be
multiplied and transferred between
bacteria in this way, via plasmids.
Figure 17.5 DNA Sequences That Move (B)
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Certain genes are present in all
organisms (universal genes); and
some universal gene segments are
present in many organisms.
This suggests that a minimal set of
DNA sequences is common to all
cells.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
Efforts to define a minimal genome
involve computer analysis of
genomes, the study of the smallest
known genome (M. genitalium), and
using transposons as mutagens.
Transposons can insert into genes at
random; the mutated bacteria are
tested for growth and survival, and
DNA is sequenced.
Figure 17.6 Using Transposon Mutagenesis to Determine the Minimal Genome
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
M. genitalium can survive in the
laboratory with only 382 functional
genes.
One goal of the research is to design
new life forms for specific purposes,
such as cleaning up oil spills.
17.2 What Have We Learned from Sequencing
Prokaryotic Genomes?
An artificial genome has been created
and inserted into bacterial cells.
The entire genome of Mycoplasma
mycoides was synthesized, then
transplanted into empty cells of
Mycoplasma capricolum.
The new cell’s genome had extra
sequences, so it was a new organism:
Mycoplasma mycoides JCV1-syn.1.0.
Figure 17.7 Synthetic Cells
Working with Data 17.1: Using Transposon Mutagenesis to Determine the Minimal Genome
• In the experiment to create a
synthetic genome and determine the
minimum set of genes necessary for
survival, transposon mutagenesis
was used with Mycoplasma
genitalium, which had the smallest
known genome.
Working with Data 17.1: Using Transposon Mutagenesis to Determine the Minimal Genome
• Growth of M. genitalium strains with
gene insertions (intragenic) was
compared with strains with insertions in
noncoding regions (intergenic).
Working with Data 17.1: Using Transposon Mutagenesis to Determine the Minimal Genome
•Question 1:
Explain these data in terms of genes
essential for growth and survival.
Are all of the genes in M. genitalium
essential for growth?
•
If not, how many are essential?
•
Why did some of the insertions in
intergenic regions prevent growth?
Working with Data 17.1: Using Transposon Mutagenesis to Determine the Minimal Genome
• Question 2:
•
If a transposon inserts into the
following regions of a gene, there might
be no effect on the phenotype.
•
Explain in each case:
a. near the 3′ end of a coding region
b. within a gene coding for rRNA
•
How does this affect your answer to
Question 1?
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
There are major differences between
eukaryotic and prokaryotic genomes:
• Eukaryotic genomes are larger and
have more protein-coding genes.
• Eukaryotic genomes have more
regulatory sequences. Greater
complexity requires more regulation.
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Much of eukaryotic DNA is noncoding,
including introns, gene control
sequences, and repeated sequences.
Eukaryotes have multiple
chromosomes; each must have an
origin of replication, a centromere,
and a telomeric sequence at each
end.
Table 17.2
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Several model organisms have been
studied extensively.
Model organisms are easy to grow and
study in a laboratory, their genetics
are well studied, and they have
characteristics that represent a larger
group of organisms.
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
• The yeast, Saccharomyces
cerevisiae:

Yeasts are single-celled
eukaryotes.

Yeasts and E. coli appear to use
about the same number of genes to
perform basic functions.

Compartmentalization of the
eukaryotic yeast cell requires it to
have many more genes.
Table 17.3
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
• The nematode, Caenorhabditis
elegans:

A millimeter-long soil roundworm.

The transparent body is made up of
about 1,000 cells, yet has complex
organ systems.

It has about 3.3 times as many
protein-coding genes as do yeasts.
Table 17.4
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
• The fruit fly, Drosophila
melanogaster:

Studies of fruit flies led to
formulation of many basic
principles of genetics. More than
2,500 mutations have been
described.

It has 10 times more cells and a
larger genome than C. elegans, but
fewer coding genes.
Figure 17.8 Functions of the Eukaryotic Genome
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
• The thale cress, Arabidopsis
thaliana:

A small plant with a small genome.

Many of the genes found in animals
have homologs in plants,
suggesting a common ancestor.

But many genes are also unique to
plants.
Table 17.5
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Rice (Oryza sativa) and a poplar tree
(Populus trichocarpa) have also been
sequenced.
Comparison of the genomes shows
many genes in common, comprising
the basic minimal plant genome.
Figure 17.9 Plant Genomes
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Eukaryotes have closely related genes
called gene families.
These arose over evolutionary time
when different copies of genes
underwent separate mutations.
Example: Genes encoding the globin
proteins all arose from a single
common ancestral gene.
Figure 17.10 The Globin Gene Family
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
During development, different members
of the globin gene family are
expressed at different times in
different tissues.
Example: Hemoglobin of the human
fetus contains γ-globin, which binds
O2 more tightly than adult hemoglobin.
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Many gene families include
nonfunctional pseudogenes (Ψ),
resulting from mutations that cause a
loss of function.
A pseudogene may simply lack a
promoter, and thus fail to be
transcribed, or a recognition site
needed for the removal of an intron.
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Eukaryotic genomes have repetitive
DNA sequences:
• Highly repetitive sequences—short
sequences (< 100 bp) repeated
thousands of times in tandem; not
transcribed.
 Short tandem repeats (STRs) of 1–
5 bp can be used in DNA
fingerprinting.
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
• Moderately repetitive sequences
are repeated 10–1,000 times.
 Includes genes for tRNAs and
rRNAs
 Single copies of the tRNA and
rRNA genes would be inadequate
to supply the large amounts of
these molecules needed by cells.
Figure 17.11 A Moderately Repetitive Sequence Codes for rRNA (Part 1)
Figure 17.11 A Moderately Repetitive Sequence Codes for rRNA (Part 2)
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
• Transposons (transposable elements)
are moderately repetitive sequences.

Three types are retrotransposons:
o
SINEs (short interspersed
elements)
o
LINEs (long interspersed elements)
o
LTRs (long terminal repeats)
Table 17.6
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Retrotransposons are transcribed into
RNA, which is a template for new
DNA. The new DNA becomes
inserted at a new location, resulting in
two copies of the transposon.
DNA transposons are excised from the
original location and become inserted
at a new location without being
replicated.
17.3 What Have We Learned from Sequencing
Eukaryotic Genomes?
Insertion of a transposon at a new
location can have important
consequences, such as mutations and
gene duplications.
They can result in shuffling the genetic
material and creating new genes.
Transposons may have played a role in
endosymbiosis.
17.4 What Are the Characteristics of the Human Genome?
Sequencing of the human genome
revealed many interesting facts:
• Protein-coding regions make up about
1.2%, or 21,000 genes.

The average gene must code for
several different proteins, and
posttranscriptional mechanisms result
in different proteins.
17.4 What Are the Characteristics of the Human Genome?
• An average gene has 27,000 base
pairs.
• All human genes have many introns.
• About half of the genome is
transposons and other repetitive
sequences.
17.4 What Are the Characteristics of the Human Genome?
• 99.5% of the genome is the same in
all people.

Variation among individuals is due to
single nucleotide polymorphisms
(SNPs), and differences in sequence
copy number from chromosomal
deletions, duplications, or
translocations.
17.4 What Are the Characteristics of the Human Genome?
• Genes are not evenly distributed over
the genome.

The Y chromosome has the fewest
genes (231); chromosome 1 has the
most (2,968).
17.4 What Are the Characteristics of the Human Genome?
Comparisons of prokaryote and
eukaryote genomes have revealed
evolutionary relationships between
genes.
Figure 17.12 Evolution of the Genome
17.4 What Are the Characteristics of the Human Genome?
The genomes of many primates have
been sequenced, and biologists are
interested in which genes make
humans unique.
Chimpanzees are our closest living
relative: they share almost 99% of our
DNA sequences.
17.4 What Are the Characteristics of the Human Genome?
DNA from the bones of Neanderthals,
who lived in Europe up to 50,000
years ago, has also been sequenced.
It is 99% identical to human DNA,
justifying classification of
Neanderthals as part of the same
genus, Homo.
17.4 What Are the Characteristics of the Human Genome?
Comparisons of human and
Neanderthal genes:
• A mutation in MC1R in Neanderthals
causes lower activity of MC1R, known
to result in fair skin and red hair.
• FOXP2, involved in vocalization, is
identical in humans and Neanderthals,
suggesting that Neanderthals were
capable of speech.
Figure 17.13 A Neanderthal Child
17.4 What Are the Characteristics of the Human Genome?
There are some distinctive “human”
DNA sequences and also distinctive
“Neanderthal” sequences.
There is some mixture of the two,
indicating that humans and
Neanderthals interbred.
17.4 What Are the Characteristics of the Human Genome?
Rapid genotyping technologies are
being used to understand the genetic
basis of diseases such as diabetes,
heart disease, and Alzheimer’s
disease.
“Haplotype maps” are used to identify
SNPs that are linked to genes
involved in disease.
17.4 What Are the Characteristics of the Human Genome?
A haplotype is a piece of chromosome
with a set of SNPs that are usually
inherited as a unit.
By comparing the haplotypes of
individuals with and without a
particular genetic disease, the loci
associated with the disease can be
identified.
Figure 17.14 SNP Genotyping and Disease
17.4 What Are the Characteristics of the Human Genome?
New technologies analyze thousands
or millions of SNPs to determine
which ones are associated with
specific diseases.
As the cost of sequencing entire
genomes decreases, SNP testing may
be superseded.
Table 17.7
17.4 What Are the Characteristics of the Human Genome?
Pharmacogenomics is the study of
how an individual’s genome affects
response to drugs or other outside
agents.
SNPs that are associated with specific
drug responses can be identified to
personalize drug treatments and
determine if a patient will respond to a
drug.
Figure 17.15 Pharmacogenomics
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Many genes encode more than one
protein.
Alternative splicing and
posttranslational modifications
increase the number of proteins that
can be derived from one gene.
But many proteins are produced only
by certain cells under specific
conditions.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Proteome: sum total of proteins
produced by an organism; it is more
complex than the genome.
Proteomics seeks to identify and
characterize all the expressed
proteins in an organism.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Two techniques are used to analyze
the proteome:
• Two-dimensional gel electrophoresis
separates proteins based on size and
electric charges.
• Mass spectrometry identifies proteins
by their atomic masses.
Figure 17.16 Proteomics
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Comparisons of eukaryotic proteomes
has revealed a common set of about
1,300 proteins that provide the basic
metabolic functions.
Figure 17.17 Proteins of the Eukaryotic Proteome
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Proteins have different functional
regions or domains.
Proteins that are unique to a particular
organism are often just unique
combinations of domains that exist in
other organisms.
This reshuffling of the genetic deck is a
key to evolution.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Gene and protein function are both
affected by the internal and external
environments of the cell.
Enzyme activities affect concentrations
of their substrates and products,
called metabolites.
As the proteome changes, so will the
abundances of metabolites.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Metabolome: quantitative description
of all of the small molecules in a cell
or organism.
• Primary metabolites—involved in
normal processes such as pathways
like glycolysis. Also includes
hormones and other signaling
molecules.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
• Secondary metabolites—often unique
to particular organisms or groups.

Examples include antibiotics made by
microbes and chemicals made by
plants for defense.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Measuring metabolites involves gas
chromatography and highperformance liquid chromatography,
which separate molecules.
Mass spectrometry and nuclear
magnetic resonance spectroscopy are
used to identify them.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
A human metabolome database has
been established and contains 6,500
metabolites.
The challenge now is to relate levels of
these substances to physiology.
17.5 What Do the New Disciplines Proteomics and
Metabolomics Reveal?
Plant metabolomics has been studied
for many years.
Tens of thousands of secondary
metabolites have been identified.
The metabolome of the model
organism Arabidopsis thaliana is now
being described.
17 Answer to Opening Question
Myostatin is a protein that inhibits
muscle growth.
In dog breeds with highly developed leg
muscles, the gene for myostatin has a
mutation that makes the protein
inactive.
In humans it may be possible to
manipulate myostatin to treat musclewasting diseases such as muscular
dystrophy.