Download Chapter 21: Genomes & Their Evolution 1. Sequencing & Analyzing Genomes

11/30/2015
Chapter 21:
Genomes & Their Evolution
1. Sequencing & Analyzing Genomes
2. How Genomes Evolve
1. Sequencing & Analyzing
Genomes
Chapter Reading – pp. 437-447
Whole Genome Shotgun Sequencing
1 Cut the DNA into
overlapping fragments short enough
for sequencing.
2 Clone the fragments
in plasmid or phage
vectors.
3 Sequence each
fragment.
4 Order the
sequences into
one overall
sequence
with computer
software.
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Bioinformatics
Bioinformatics refers to application of statistics and
computer analysis to DNA, protein sequence data.
• computer
analysis can
identify protein
coding regions
in DNA,
determine amino
acid sequences,
compare
sequences
among species,
etc…
Systems Level
Analysis
Translation and
ribosomal functions
Glutamate
biosynthesis
Mitochondrial
functions
Vesicle
fusion
RNA processing
Peroxisomal
functions
Transcription
and chromatinrelated functions
Metabolism
and amino acid
biosynthesis
Nuclearcytoplasmic
transport
Secretion
and vesicle
transport
Nuclear migration
and protein
degradation
Mitosis
DNA replication
and repair
Cell polarity and
morphogenesis
Protein folding,
glycosylation, and
cell wall biosynthesis
Serinerelated
biosynthesis
Amino acid
permease pathway
Computer
analysis can
predict
protein/protein
interactions
and thus
protein
function.
Relative Genome
Size
Genome size and gene
number do not correlate
at all with organism
complexity.
• alternative splicing of genes
and the repertoire of noncoding RNAs (e.g., miRNA)
may be a better indicator
of “sophistication” or
complexity in a species
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Types of Human DNA Elements
Most of the
human genome
(and that of
many other
species) does
not code for any
obvious gene
products and
has a function
that is as yet
unclear.
Repetitive DNA Elements
Much of the human genome consists of
repetitive DNA sequences that are thought to
ultimately be of viral origin.
Transposable Elements
• DNA segments that are duplicated and distributed
throughout the genome
Alu elements
• repetitive DNA sequences containing the Alu I
restriction enzyme sites
Short tandem repeats (STRs)
• very short sequences repeated over and over
Transposable Elements
Barbara McClintock proposed the concept of
“jumping genes” in the 1950s based on her
studies of corn which was not taken seriously.
Much later the
existence of
transposable
elements that could
“jump” in the
genome validated
her observations.
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Transposons
Transposon
DNA of
genome
New copy of
transposon
Transposon
is copied
Insertion
Mobile transposon
Mobile DNA elements that can be copied & inserted
Elsewhere in the genome.
• the transposon encodes the enzyme transposase which can
copy transposon sequence and randomly insert elsewhere
Retrotransposons
Retrotransposons are much like transposons except that
they encode reverse transcriptase and have an
RNA intermediate in the process.
New copy of
Retrotransposon
retrotransposon
Formation of a
single-stranded
RNA intermediate
RNA
Insertion
Reverse
transcriptase
Multigene Families
Many genes are actually part of a group or
cluster of similar genes referred to as a
“multigene family”.
• 2 or more genes with nearly identical or very
similar sequences
• thought to have arisen due to gene duplication
and subsequent mutation
• members of a multigene family are typically
similar in function as well as sequence
• arrangement of genes in multigene families also
provides evidence of similar origins
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DNA
RNA transcripts
Nontranscribed
spacer
Transcription unit
DNA
18S
28S
5.8S
rRNA
5.8S
28S
18S
(a) Part of the ribosomal RNA gene family
-Globin
-Globin
Heme
-Globin gene family
-Globin gene family
Chromosome 11
Chromosome 16

Embryo
 
2

1
2
1

Fetus
and adult

Embryo
G
A

Fetus


Adult
(b) The human -globin and -globin gene families
2. How Genomes Evolve
Chapter Reading – pp. 448-458
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Rearrangement of Genomes
Genomes can undergo a number of large-scale
changes that can lead to significant changes in
genetic structure and in gene products:
Chromosomal rearrangement
• breaking and recombining of pieces of diff. chrom.
Transposition of mobile DNA elements
• sequences that can move around the genome
Gene duplication
• duplication of gene sequences
Exon shuffling
• combining of exons from different genes
Changes in Chromosome Structure
Human chromosome 2 is clearly a combination of
chimpanzee chromosomes 12 & 13.
Human
chromosome 2
Chimpanzee
chromosomes
Telomere
sequences
Centromere
sequences
Telomere-like
sequences
12
Human
chromosome 16
Centromere-like
sequences
13
(a) Human and chimpanzee chromosomes
Blocks of genes in mice
and humans have
remained intact though
they are distributed
differently among
chromosomes.
Mouse
chromosomes
7
8
16
17
(b) Human and mouse chromosomes
Evolution of Novel Genes
Genes encoding proteins with entirely new
functions can arise by:
1) Duplication of existing gene followed by
mutation producing distinct gene product
• the 2 genes will share significant homology
however may have very different functions
(e.g., lysozyme and -lactalbumin)
2) Exon shuffling
• errors in meiotic recombination or transposition can
cause the addition or loss of exons from similar or
very different genes
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Gene Duplication & Crossing Over
Nonsister Gene
chromatids
Transposable
element
Misalignment of
similar DNA
sequences
during meiotic
crossing over
can result in
chromosomes
with duplicated
(or missing)
regions of DNA.
Crossover
point
Incorrect pairing
of two homologs
during meiosis
and
Duplication followed by Mutation
lysozyme vs -lactalbumin
Model for Globin Gene Duplication
The globin gene families show evidence of duplication.
Ancestral globin gene
Evolutionary time
Duplication of
ancestral gene
Mutation in
both copies

Transposition to
different chromosomes
Further duplications
and mutations






      2 1  
2
1
-Globin gene family
on chromosome 16



G

A



-Globin gene family
on chromosome 11
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Globin Gene Comparison
Over time apparently duplicated globin genes
diverged via mutation into similar yet distinct
proteins with similar yet unique functions.
Exon Shuffling
EGF
EGF
EGF
EGF
Epidermal growth
factor gene with multiple
EGF exons
F
F
F
Exon
shuffling
Exon
duplication
F
Fibronectin gene with multiple
“finger” exons
F
EGF
K
K
K
Exon
shuffling
Plasminogen gene with a
“kringle” exon
Portions of ancestral genes
TPA gene as it exists today
Comparing Genomes
Bacteria
Most recent
common
ancestor
of all living
things
Eukarya
Archaea
4
1
3
2
Billions of years ago
0
Chimpanzee
Human
Mouse
70
60
50
40
30
20
10
0
Sequence
homology and
genome structure
reflect
evolutionary
relatedness:
• degree of
differences in
gene sequences,
chromosome &
gene structures
allow estimation
of time since a
common ancestor
Millions of years ago
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Gene Knockout Experiments
For the last 20 or so years we have had the
technology to disrupt or “knockout” genetic
alleles in mice to assess gene function:
1) mouse embryonic stems cells (ES cells) are
genetically altered in vitro
• specific gene is targeted for disruption and ES cells
with disrupted gene are selected for
2) disrupted ES cells added to early embryos which are
implanted into surrogate mothers
3) offspring are screened for “knockout” allele
4) positive offspring are mated to produce homozygous
knockouts for analysis of gene function
A Gene Knockout
EXPERIMENT
Wild type: two normal
copies of FOXP2
Heterozygote: one
copy of FOXP2
disrupted
Homozygote: both
copies of FOXP2
disrupted
Experiment 1: Researchers cut thin sections of brain and stained
them with reagents that allow visualization of brain anatomy in a
UV fluorescence microscope.
mice don’t
“whistle” (i.e., vocalize)
when stressed*.
Foxp2-/-
RESULTS
Experiment 2
Number of whistles
Experiment 1
Experiment 2: Researchers separated
each newborn pup from its mother
and recorded the number of
ultrasonic whistles produced by the
pup.
Wild type
Heterozygote
400
300
200
100
Homozygote
*FOXP2 must be necessary for assoc. brain develop.
Homeotic
Genes
Homeotic genes were
first discovered in
Drosophila to specify
body segments.
Homologous genes
arranged in same
fashion have been
identified in mammals
and other animals as
well as yeast & plants
• Hox gene clusters or
families in animals
(No
whistles)
0
Wild
type
Heterozygote
Homozygote
Adult
fruit fly
Homeotic genes
encode TFs
Fruit fly embryo
(10 hours)
Fly chromosome
Mouse
chromosomes
Mouse embryo
(12 days)
Adult mouse
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Differential Hox Gene Expression
Thorax
Genital
segments
Thorax
Abdomen
Abdomen
Differences in
Hox gene
expression result
in different
specification of
body segments
and thus
different overall
body plans
among species!
Key Terms for Chapter 21
• 3-step genome vs whole genome shotgun seq.
• bioinformatics, genomics, proteomics
• transposable elements: transposons , transposase,
retrotransposons
• gene duplication, exon shuffling
• gene knockouts, knockout mice, ES cells
• homeotic genes, Hox genes
Relevant
Chapter
Questions
1-6
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