Download Model organisms and mutants

Course Overview
Model organisms and mutants
http://www.erin.utoronto.ca/~w3bio/bio207/index.htm
March 6
Gene mutation
• Gene mutation: a gene can mutate from one
allelic form to another
– Forward mutation:
• A+ to a
• D+ to D
– Reverse mutation:
• a to A+
• D to D+
Outline
Week
1
2
3
4
5
6
7
8
9
10
11
12
Topic
Course objectives and Introduction to genetics
Human Pedigrees
Patterns of Inheritance: sex-linkage
Chromosomal basis of inheritance
Changes in chromosome number
Gene Mapping
Gene to Phenotype
Modified Mendelian ratios
Model organisms and mutants
Genetics of Plant Development (Arabidopsis)
Genetics of Animal Development (Drosophila)
Behaviour Genetics/Quantitative genetics
Chapter
Ch. 1 & Ch. 2
Ch. 2
Ch. 2
Ch. 3
Ch. 15
Ch. 4
Ch. 6
Ch. 6
Ch. 6 &Ch. 16
Ch. 18
Ch. 18
Ch. 16 + papers
Model organisms
• Selected by researchers based on some
feature that renders it particularly useful
for studying the genetic process of interest
to that researcher.
• Each model organism usually has a
database and a community of researchers
• Mutations can be used for two purposes:
– To study the process of mutation itself
– To permit the dissection of a biological function
Text p. 14-15 & 707-23
1
Drosophila melanogaster
• Database: FlyBase < http://flybase.org>
• Mutations in fruit flies
Drosophila melanogaster
• Life cycle of a fruit fly
– E.g. Vestigial wings (vg)
Drosophila melanogaster
• Lethal genes: some alleles of a gene are lethal
in the homozygous form. This could be the
homozygous recessive or the homozygous
dominant genotypes that are lethal. This
changes the ratios.
– For example with the Cy (curly) gene in Drosophila it
is lethal as a homozygous dominant Cy/Cy so the
phenotypic ratio you get from a hybrid cross is 2:1.
When calculating the ratio you do it from the progeny
that live (these are the only ones you can score!).
Lethality in Drosophila
• Cy is a homozygous
lethal mutation
• Cross two
heterozygotes
– Cy/+ X Cy/+
• Cy /+ X Cy/+
–
–
–
–
Cy/Cy
Cy/+
+/Cy
+/+
dies
lives
lives
lives
Curly
Curly
wildtype
• Get a 2:1 ratio of
Curly to wildtype
2
ClB chromosome
• In 1928 Herman Muller devised a method
of searching for any lethal mutation on the
X-chromosome (chromosome 1 in
Drosophila)
• He constructed a chromosome called ClB
which carries an inversion a cross-over
suppressor (C), a lethal (l) and a dominant
mutation Bar-eye (B) marker.
3
Balancers
• Since Muller constructed the ClB chromosome many
other chromosomes bearing inversions have been
constructed
• Chromosomes bearing multiple inversions are used to
suppress crossing over and are often marked with
recessive lethal mutations
• Only offspring that are heterozygous for the inversion
AND the lethal mutation survive
• Balancers e.g.:
– FM7 First chromosome balancer
– SM6 Second chromosome balancer marked with Cy
– TM3 Third chromosome balancer marked with Stubble
Deletion mapping
• Deletions change
dominance
relationships
• Pseudodominance:
recessive alleles
appear dominant over
a deletion
– Eg pn shows
pseudodominance with
deletion 264-38
• Recessive lethal
mutations are
maintained over a
balancer chromosome
Mutations and screens
• Making mutations in
Drosophila melanogaster
• Enhancer trap screens
• Transposable elements
are used as mutagens
because as they “hop”
they disrupt the integrity
of a gene. The gene is
identified by transposon
tagging (see chapter 13
p438)
• Reverse genetics also
uses P-elements to insert
DNA into the fly
4
Mutations and screens
• Making mutations in
Drosophila melanogaster
• Enhancer trap screens
• Transposable elements
are used as mutagens
because as they “hop”
they disrupt the integrity
of a gene. The gene is
identified by transposon
tagging (see chapter 13
p438)
• Reverse genetics also
uses P-elements to insert
DNA into the fly
Mus musculus
• Database: MGI <http://www.informatics.jax.org>
• Mouse coat colour genetics
5
Lethal genes
• Lethal genes: mutant alleles capable of
causing the death of the organism
• Mouse coat colour mutation yellow is
homozygous lethal allele of Agouti (A)
– yellow X yellow
Lethal genes
• AY / A
–¼
–½
–¼
x
AY / A
AY / AY
AY / A
A/A
lethal
yellow
wildtype (Agouti, Black)
gave
• 2/3 yellow, 1/3 wildtype progeny
– Describe with symbols what you think is
happening
Mice
• Mus Musculus
• Specific fragments of
DNA can be inserted
into the genome of
mice
– E.g. the jellyfish gene
for green fluorescent
protein
Text ch. 11 p.374
6
Neurospora crassa
• N. crassa
• Growth mutants
• Database:
http://www.mips.biochem.mpg.de/proj/neurospora/)
• Genome sequenced in 2003
Text ch. 21 p. 700
Neurospora “sex”
• Mating type is a simple
form of sex
• Mating types are
determined by two alleles
of a single gene: MAT-A
and MAT-a.
• Crosses between the two
haploid mating types
produces diploid
meiocytes then
ascospores
Forward genetics
• Forward genetics
often uses genetic
selection protocols in
which only the
desired mutant
phenotype survives
– E.g. nutritional
mutations in N. crassa.
7
Testing strains
• 20 progeny of a cross
ad· leu+ X ad+· leu
• What is the genotype of culture 8?
• What is the genotype of culture 9?
8
Testing strains
• 20 progeny of a cross ad· leu+ X ad+· leu
• What is the genotype of culture 8?
– ad· leu+
• What is the genotype of culture 9?
– ad· leu
Neurospora crassa
• Forward genetics
often uses genetic
screens protocols in
which the desired
mutant phenotype is
selected from among
a large number of
individuals
– E.g. growth mutants in
Neurospora
Reverse genetics
• In Neurospora a useful mechanism for
targeting mutations to a specific gene is
Repeat-Induced Point mutation (RIP)
• A strain with an ectopically induced
wildtype transgene is crossed
• In the resulting strains the transgene has
many mutations (GC· AT transitions)
9
Caenorhabditis elegans
C. elegans
•
•
•
•
Database: WormBase http://www.wormbase.org
Nematode, a soil-dwelling roundworm
Hermaphrodite
Transparency makes it easy to for screen for mutations
Synthetic lethals
• Synthetic lethals are formed when two
viable single mutants are intercrossed and
the resulting double mutant is lethal
10
Reverse genetics: RNAi
• RNAi inactivates the
homologous gene
• The introduced double
stranded RNA is cut into
many segments
(interfering RNAs)
• RNAs complementary to
the interfering RNAs are
degraded
• In the absence of mRNAs
the organism expresses
the mutant phenotype
Text p. 534
Arabidopsis thaliana
11
Arabidopsis thaliana
• Arabidopsis database: TAIR
http://www.arabidopsis.org
• Arabidopsis and the genetics of
development
Zebra fish
• Danio rerio (Zebra fish) are becoming a genetic model
for vertebrate development and neurobiology
• Database: ZFIN http://zfin.org
• Visible mutants
– E.g. mutants for haematopoesis and stripe pattern
Developmental mutants in zebra
fish
12
Introduction to genetic analysis
Griffiths, A., Wessler, S.R., Lewontin,R.C., Gelbart, W.M.,Suzuki, D.T.
and Miller, J.H.
Eighth Edition, W.H. Freeman and Company NY
• Part I Transmission genetic analysis
–
–
–
–
Chapter 1: all questions p. 24-26
Chapter 2: all the questions p. 62-72
Chapter 3: questions #1-12,18,19, 22, 25-27, 29, 30, 32, 40-42.
Chapter 4: sections 4.1- 4.4 and 4.6, questions # 1-4, 6-13,15-22,24-43.
• Part II The relationship of DNA and phenotype
– Chapter 6: questions# 1,3-5,7,9,11,14,18,19,21,23,24,
26,29,32,33,35,39,41,44,45,47,49,51,53,55,63-66.
• Part IV The nature of heritable change
– Chapter 15: sections 15.1 and 15.3; questions #1-3,11-13,19,21,22, 32, 38, 52.
• Part V From Genes to processes
– Chapter 16: questions# 1,17,19,21,22,25,28,
30,31,32,35,39; (not required to know yeast, Aspergillus
and E.coli information).
.
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