Download Mouse Genetics

An Overview of Mouse Genetics
Reference Material
 Mouse Genetics, Concepts and
Applications. 1995. Silver, L.M.
Oxford University Press, New York,
NY, 1995, pp 1-362.

Genetic Variants and Strains of the
Laboratory Mouse, 3rd Edition.
1996. Edited by M. F. Lyon, S.
Rastan and S.D.M. Brown. Oxford
University Press,New York, NY, 2
Volumes, pp. 1-1807.
Reference Material
(Out of Print; Often in Libraries)
 The Mouse in Biomedical Research. 1981.
Edited by H.L. Foster, J.D. Small
and J.G. Fox.Academic Press, New
York,NY.4 Volumes.
 Inbred Strains in Biomedical Research.
1979. Festing, M.F.W. Oxford
University Press, New York, NY.

Origins of Inbred Mice. 1978. Morse
III,H.C. Academic Press, New York,
NY.
Genetics I: From Fancy Mice to Mendel
 Brief History of the Laboratory Mouse
 Details and Definitions
 Genetic Resources
 Gene Mapping
 Annotating the Mouse Genome
Origin of the House Mouse
Geographical Ranges of Subspecies
Evolutionary Relationships Among Commonly
Used Model Organisms
Mouse as a Model System
 Small size: 25-40g (2,000-3000 fold
lighter than an average human)
 Short gestation time: ~10 weeks from
being born to giving birth
 Breed well: ~5-10 pups/litter and an
immediate postpartum estrus
 Fathers do not harm young
 Vaginal plug: timed pregnancies
Mouse Genome
 3 x 109 base pairs

~1450 cM

~2Mb/cM

~25,000 genes

Organized into 19 autosomes and the X
and Y Chromosome

Chromosomes are acrocentric and show a
continual decrease in size
Genetic Resources
 Outbred Stocks

Inbred Strains and Substrains

F1 Hybrids

Mutant Strains
Outbred Stocks
 Genetically undefined

Bred to maintain a maximal level of
heterozygosity in all animals

Long life spans, high disease resistance,
large and frequent litters, low
neonatal mortality, rapid growth,
large size, CHEAP

Examples: CD1, Swiss Webster, ICR, NIH

Useful in experiments where genotype does
NOT matter (biochemical purification, stud
males)

Should NEVER be used in any breeding
experiment because of the genetic
uncertainty
Inbred Strains
 All members of a strain are genetically
identical
 Bred by strict brother X sister matings
for at least 20 generations (Note:
~3-4 generations/year; at least 5
years)
Consequences of Inbreeding up to 20
Generations
Beck J. A. Nat. Genet. 2000;21:23-25
FVB/NJ
AKR/J
NOD/LtJ
BALB/cWt
MRL/MpJ
AKR/J
Lymphatic leukemia
BALB/cWt
Y- chromosome nondisjuction
FVB/NJ
Male pronucleus in
fertilized egg is
large
MRL/MpJ
Lymphoproliferation
NOD/LtJ
Diabetes
Substrains
Simpson E. M. Nat. Genet. 1997;16:19-27
Distribution of SNPs and Haplotypes on Chromosome 4
F1 Hybrid Mice

Produced by crossing mice of two
different inbred strains

Can be repeatedly produced as long as
the parental strains exist

Heterozygous at all loci at which the
parental strains differ

Genetically and phenotypically uniform

Do not breed true (F2 generation mice
undergo recombination)
Desirable Features and Uses of F1 Hybrid
Mice

Hybrid vigor: Increased disease resistance,
better survival under stress, greater
longevity and larger litters than inbred
mice

Useful for carrying deleterious mutations,
in radiation research, and for
bioassays of pathogens, hormones,
drugs, etc……….

Accept transplants of tissue (tumors,
skin, ovaries) from mice of either
parental strain.
Some Common F1 Hybrid Strains
bgJ
white
nose
Aw-J
c2J
me
“A Mutation”
 What is it?: A variation from wild-type at a
single locus

Mode of origin: Spontaneous or induced
(Chemicals, Radiation, Gene Modification)

Mode of inheritance: dominant,
semidominant, recessive (or not
heritable)

Penetrance: complete or incomplete

Interaction with other mutations
Some Practical Information about
Mutations
 Mouse Gene Symbols: always italicized (or
underlined), assigned by the
International Mouse Genetics
Nomenclature Committee, change
often

Example: Dom; rec

Serialization: Dom1, Dom2; rec1, rec2

Alleles of the Same Locus: Dom1a, Dom1b

Special rules for pseudogenes, transgenics, etc..
A Wealth of Information at….
 The Jackson Laboratory Home
Page:http://www.jax.org
Mutant Strains
 Coisogenic/Isogenic: mutation arose on an
inbred strain and is still maintained
on the strain of origin (C57BL/6J-M)
 Congenic: constructed by transferring a
chromosomal segment carrying a
locus, phenotypic trait, or mutation
of interest from one strain to
another by 10 or more successive
backcross matings (B6.C-H-2d)
Genomic Homogeneity and Heterogeneity
During the Creation of a Congenic Strain
Average Length of the Differential
Chromosomal Segment During the Creation
of a Congenic Strain
Why Make Congenic Strains?
 Evaluate phenotype of a mutation (single
gene trait) on one or more well
defined genetic backgrounds

Can be very useful in the analysis of
complex traits
The Egfr (ko/ko) ExamplePhenotype Dependent on Genetic
Background

Peri-implantation lethal on a CF-1
background due to degeneration of the
inner cell mass

Mid-gestation lethal on a 129/Sv
background due to placental defects

Survive up to weaning with abnormalities
in many organs on a CD-1 background
 Treadgill et al., Science:269, 230-240,
1995.
Markel P. Nat. Genet. 1997;17:280-284
Consomic Strains
 Very similar to congenic strains except that
selection is for a whole chromosome
instead of a chromosomal segment

A set of consomic strains with each A/J
chromosome on a C57BL/6J background
have been constructed

Particularly useful for the analysis
quantitative trait loci (QTL’s) and
modifier loci

Singer et al., Science: 304, 445-448, 2004
Recombinant Inbred Strains
Recombinant Congenic (RC) Strains
 Similar to RI strains, initial cross is between
two distinct inbred strains

Next two generations are made by
backcrossing without selection to one
of the parental stains

Next 14 generations of brother x sister
matings are performed

Mosaic genomes skewed in the direction of
the backcross parent (7/8 versus 1/8)
Gene Mapping-Definitions
 Locus - a DNA segment that is
distinguishable in some way by some
form of genetic analysis (gene,
anonymous DNA, etc…)
 Genetic map - a representation of the
distribution of a set of loci
within a genome (linkage,
chromosomal, and physical)
Why Map Genes
 Facilitate moving from disease phenotype
to cloning the causative gene(s)
(positional cloning)
 Can provide function for a recently cloned
gene
 Can be used to dissect out the heritable
and nonheritable components of
complex traits

Comparative genetics
Linkage Maps
 Also known as recombination (meiotic)
maps
 Can only be constructed for loci that
occur in two or more heritable forms
or alleles
 Are generated by counting the number of
offspring that receive either the
parental or recombinant allele
combinations at two or more loci
Where Genetic Recombinants Come From
Independent Assortment of Alleles from
Unlinked Loci
Non-independent Assortment of Alleles
from Linked Loci
Gene Order
White, circling, not pudgy
Pigmented, circling, not pudgy
Pigmented, not circling, pudgy
Gene order pu - c/Tyr - sh1 is derived from this
multipoint cross
by parsimony (I.e., that in which there are few
or not double-crossover events)
A more popular way of looking at it:
nonrecombinant
recombinant
pu
c, Tyr
sh1
# mice
46
Heterozygous, or “b” allele
inherited from F1 parent
51
1
1
1
Homozygous, or “a” allele
inherited from pigmented parent
Live Linkage Map of the Mouse
10th International Congress of Genetics
Montreal, Canada. 1958.
Non-Meiotic Mapping Methods
 Somatic cell hybrids
 Radiation hybrids
 in situ hybridization
The Ultimate Physical Map
 The sequence of the mouse genome

Public draft sequence (~96% of the euchromatic
genome of C57BL/6J excluding the Y
chromosome) released in December
2002

Sanger Center is completing the sequence
of chromosomes 2, 4, 11, and X

Page lab has sequenced part of the mouse Y

The rest?
Functional Annotation of the Mouse Genome
 Need: One or more independent mutations
in every gene
 Approach: Genotype to phenotype
 Approach: Phenotype to Genotype
Genotype to Phenotype
 Targeted mutations in each gene
 Gene trap mutagenesis
 Chemical mutagenesis of ES cells
 Ultimately, must determine the
phenotypic affect of the mutation,
in vivo
Targeted mutations
 Homologous recombination in ES cells
 Advantages: can create exactly what you
want;conditional or null allele, temporal
regulation, point mutations, small deletions,
etc.; tend to know a lot about the gene
which can help with interpreting phenotype
 Disadvantages: time-consuming and very
expensive if all ~25,000 genes are to be
mutated; must make assumptions about
important functional domains
Targeted mutations
the future?
 Development of simpler/faster methods for
construct building-”recombineering” and high
throughput analysis
 The Sanger Plan--generate 10,000 - 15,000
targeted mutations (probably conditional, not null) in
a five year period; A FEW THOUSAND IN THE
GERMLINE
 Part of a larger European effort that will include a
collection of null alleles, total exact numbers are
unclear
 US effort--discussion stage
Gene Trap Mutagenesis
 A form of insertional mutagenesis in ES cells
 Various vectors have been developed but all are
designed to report the expression of the
endogenous gene at the site of integration
and provide a DNA tag for rapid
identification of the disrupted gene
 Lexicon:60% coverage of the mouse genome from
200,000 sequence tags
 International Gene Trap Consortium (IGTC):32%
coverage in 27,000 tags (variety of vectors
may overcome insertion site preference);
20% do not overlap with Lexicon
Gene Trap Mutagenesis
 Advantages: IGTC clones are available without
restriction (see Nature Genet. 36, 543545, 2004); high throughput (1 new gene
added every 35 tags); combination of
vectors helps overcome integration site
preference; like targeted mutations,
insertions can be archived as ES cells,
which is inexpensive
 Disadvantages: Unclear what you have:null,
hypomorphic, or neomorphic allele; most of
the existing insertions have not been
examined for phenotype, in vivo
Gene Trap Mutagenesis
the Future?
 IGTC plans to add additional tags ( until ~ saturation)) to
expand the pool of tagged genes freely available
 IGTC plans on additional refinement of the
technology (new vectors, post-insertional
modification of trapped loci to create
additional/desired alleles of a tagged gene)
 Likely will be up to individual investigators to put the
mutations in the germ line and phenotype mice
 Toronto Component of the IGTC has done phenotyping on
some of the animals
Chemical Mutagenesis of ES Cells
 Induce mutations in ES cells with ENU
 A powerful chemical mutagen that primarily
induces point mutations or small deletions
 Correlate mutant phenotype with the affected gene
Chemical Mutagenesis in ES Cells
 Advantages: can directly screen for mutations in
gene of interest and then make mice from
the mutant cells;can create an allelic series
of mutations, can also vary the mutagen to
induce different types of lesions
 Disadvantages: like gene traps, it takes some work
to determine what type of mutation you
have-null or hypomorphic
Phenotype to Genotype
 Induce and select for a mutant phenotype
 Map the position of the mutation-genome scan
 Positionally clone the mutation
 Correlate mutant phenotype with the affected gene
ENU Mutagenesis, in vivo
 A powerful chemical mutagen of spermatogonial
stem cells
 Primarily induces point mutations or small deletions
 Can select for dominant, semidominant or recessive
mutations; mutations can be gain or partial
or complete loss of function
 One mutation/locus in ~600 - 1000 gametes
Enu Mutagenesis Projects
 International effort-large scale, genome wide
 Centers in England, Germany, Japan, Toronto
(http://www.mut.har.mrc.ac.uk)
 Jax, Northwestern, Oak Ridge, Baylor and others
 A few smaller genome wide efforts, as well-Sloan
Kettering
ENU Mutagenesis
 Advantages-always have a phenotype, can select
for particular organ system/stage of
development/tissue type; can get
hypomorphic alleles (new alleles of existing
mutations)
 Disadvantages-time consuming especially for
recessive screens; works best with robust
phenotypes;must go through a round of
mapping but positional cloning is greatly
simplified with the sequence of the
genome;archiving and distributing the
mutation is much more work than with ES
cells
Issues for both Approaches
 Phenotyping
 Databases
 Distribution