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Effective Colony Management
Mouse 101 Lecture
5/22/2018
Wendy du Bois, Biologist
NCI/CCR/LCBG
Why Use Mice as a ModelO Biologically very similar to humans: 95% shared
genes, very similar physiological systems, get similar diseases for many of the same genetic reasons
O Can manipulate the genome directly and model specific human diseases
O Inbred mice are available that are genetically identical to each other. Increases accuracy and reproducibility in experiments
O Accelerated life span: 1 year in a mouse= 30 years in a human
O Have been used in research for more than 100 years. Very well understood and described
O Cost effective- small, reproduce quickly, easy to handle and transport
Silver LM 1995
Mouse Lifespan
O 1-3 years
O Neonate: birth to wean (~21 days), begin eating solid food at 12-14 days
O Sexual maturity at 5-8 weeks (5-7 for males)
O Adult size at 8-10 weeks
O Geriatric at 18+ months
O Gestation: 18.5-21 days
O Litter size: 2-12+ pups
O Weaning age: 17-28 days
Breeding Objectives
O Maximize quality- healthy offspring of the desired genotype
O Minimize cost- avoid excess pups (genotype early), recognize and replace poor breeders
O Maximize pup yield to meet research goals
O Use Punnett square calculators to help choose the most effective breeding scheme
http://www.silverfallchinchilla.com/genetics/PunnetCalculator.asp
Setting Up Breeder Pairs
O Sexual maturity 5-8 weeks- set up 6-8 weeks*
O Breeders should be replaced every 7-8 months
(optimal breeding age 2-10 months)
O Replace breeders if they are not productive
1. no litter 60 days after mating (90 for
mutant strains)
2. 60 days since last litter
3. 2-3 litters born with no weaned pups
4. can try rotating males
Mice can be set up as either:
O Monogamous pairs: one male with one female
O This takes advantage of the post partum estrous cycle and can reduce the chance of fighting
O Takes up space, so higher cost. Older litter has to be weaned before new litter is born. Can be a problem with difficult to breed strains
O Harem Mating: 1 male with 2-3 females
O Pregnant females must be separated to prevent over crowding
O Can be difficult to track progeny
O Helpful if you need to produce larger numbers or if you have a limited number of males
Genetic ModificationTransgenics, Knock-ins, Knock-0uts
O Things to consider:
O May lead to embryonic lethality-
common in homozygotes
O Infertility
O May lead to poor mammary
function
O Disease: tumor development,
neurodegeration
Generating a New Mouse ColonyO Generally a good idea to start with 2-4 breeder pairs
if possible.
O How will you be identifying the mice you want to use?
O Genotype? PCR or Southern
O Phenotype?
O Breeding in the case of x-linked genes
O Progeny testing
Mouse IdentificationO Neonates:
O Toe clip (up to 7 days)
O Ear notch
O Tattoo
Weanlings/Adults: Ear Notch: inexpensive, can use tissue for genotyping
can tear and heal
Ear Tag: relatively inexpensive, unique numbering
can fall out, infection, difficult to read, mice have
to be older
O Tattoo: permanent, easy to read
O difficult to do, time consuming, can fade
O Microchips: permanent, unlimited numbers, can
also record biometric information/expensive
GenotypingO Can use either small piece of tail or a piece of ear
tissue
O Various kits available for extracting DNA
O Phenol/Chloroform and other precipitation methods
O HotSHOT:
O Alkaline lysis reagent
O Heat at 95 degrees for 10 to 60 minutes(30 minutes optimal)
O Cool to 4
O Add TrisHCl neutralization buffer
O Use 1-3ul directly
O Store at 4 to -20
Be alert to changes in productivity
O Things that can influence breeding:
O Hybrid vigor
O Post natal effects
O Parental behavior
O Genetic background
O Diet- high fat vs low fat
O If your pups are being cannibalized, things to consider:
O Is it the mom’s 1st litter?
O Is the male aggressive?
O Is there a 2nd litter
Fostering At Risk LittersO Necessary if mother dies or is neglectful,
has poor milk production (look for milk spot) or if her litter is too large.
1. Use a mouse with a different coat color.
2. Choose a female that has already successfully weaned a litter.
3. Be sure she has a litter of pups that are roughly the same age
4. Sac some of her pups if necessary
1. Move the foster mom to a clean cage
2. Using clean gloves add the pups to her dirty cage and mix the pups in with her litter
3. Wait a few minutes, then return her to the cage. You can add a treat to distract her.
4. Watch from afar- she should begin to care for the pups within one hour
5. DO NOT forget to record what you have done on the cage card!!
Environment Is Crucial to a Productive Colony
O Temperature: 68-79 degrees
O Light cycle and intensity
O Noise/vibration
O Air pressure and humidity
O Odors
O Can be sensitive to changes in caretaker staff
O Season
O Enrichment
Animal Health Issues
Signs that your mouse is sick:
❖ scruffy coat
❖ hunched, sunken at hips
❖ weight loss-malocclusion?
❖ labored breathing
❖ skin lesions
❖ eye or nasal discharge
❖ abnormal behavior
❖ dystocia/retained pups
Pedigree Record Keeping
O Critical to animal studies
O Maintain a pedigree book or database
Include:
• History of strain/genetic background
• Name changes
• Parentage information
• Generation number (N=number of times mated to the background strain. F= filial or number of times brother sister mated. p= cryopreserved)N6F4p
• Genotypes/tissue numbers
• Dates of mating
• Date cage retired
• Info about quality control tissues (what kind, location)
• Litter information
Mouse Colony Management Software Options
O Excel spreadsheets
O JaxColonyManagementSystem (JCMS)
http://colonymanagement.jax.org
free, can print cards, manage experiments, excellent tutorials
O Softmouse
https://www.softmouse.net
Has a free version, depends on size of lab/colony, calendar with email reminders about breeding etc., can print cards
O Mousecolony
www.mousecolony.com
~$550/user
FileMakerPro
Cage CardsO Should include:
O Protocol number
O Investigator name
O Strain name
O DNA info (id number/genotye)
O Sex
O Number of mice in cage
O Date of Birth
Rockvilleprinting.com
Breeder CardsO Can get important
information from
breeder cards:
O Date female found
pregnant
O Date litter born
O Litter size at birth
O Number of pups
weaned
O Ratio of m:f at wean
O Keep ALL Cards!
O Use different
colors for
different strains
O Can make
specialized
cards-timed
pregs etc.
Long Term ConsiderationsO Maintain founder stocks
O Check for Genetic Quality control
O Keep reference tissues at a set number of
generations (i.e. every time you advance an
N-generation or every 5 F’s)
O Consider having strains tested by an outside
source for “purity”
O Jackson Labs ($179/sample)
O Charles River
O Dartmouse ($149/sample, 5307 available SNPs)
Dartmouse QC Report
Consider Cryopreservation
O Gets mice you are not actively using off the
shelf- saves $$$$
O Provides a way to eliminate pathogens
O Helps to prevent genetic drift/mutations
O Disaster prevention-fire, flood, disease
O Insurance and peace of mind:
O Development and basic phenotyping of a typical strain is 2-3yrs and >$100,000
jax.org
Methods of Cryopreservation
O Embryo: $337/strain, usually
have to provide 4-6 males which
are mated to comercially
available inbred females
O Ovary: $644/strain, useful for
strains with “weak” embryos
O Sperm: $1084-1455/strain
O Jax also has a “do it yourself”
sperm freezing kit- $3500 for 3
strains, includes shipping, QA
and storage for 3 years.
Consider Donating Your Strain
O NCI mouse repository: funded by NCI for
mouse cancer models. Strains made
available to all members of the scientific
community (required by the NIH).
O Jackson Labs: you pay to ship, strains are
rederived and made available to other
scientists.
On Line Resources Available
O NCI LASP: ncifrederick.cancer.gov
O Jackson Labs Mouse Database: www.jax.org
O Mouse genome informatics:
www.informatics.jax.org (genome database, gene
expression, tumor biology, polymorphisms)
O Jax Phenome Database:
www.phenome.jax.org (collaborative, standardized collection of measured data, includes baseline phenotype data)
O International Mouse Strain Resource:www.findmice.org (searchable online database of mouse strains. Goal is to assist the international scientific community in locating and obtaining mouse resources for research)
Services Available at NIH
NCI/Frederick/LASP:
A. Colony expansion
B. Speed congenics
C. Generation of transgenic and gene targeted mice
D. Small animal imaging (xray, ct, pet, optical bioluminescence and fluorescence)
E. Pathology/help with study design
F. Embryology
G. Blood chemistry analysis/hematology
H. Technical regulations, guidelines, policies
OthersO Division of Veterinary Resources
(DVR)
O bacteriology
O diagnostic support/ pathology
O health surveillance/monitoring
O nutrition
O pharmacy
O behaviorist
O irradiation
O NHLBI Phenotyping Core
O cardiovascular phenotyping
O metabolic phenotyping
O neuromuscular
O behavioral
O exercise physiology
O advanced imaging
The End!
Jackson Labs www.jax.org
Fox, J. et al. The Mouse in Biomedical Research
Silver, L.M. (1995). Mouse Genetics
https://ncifrederick.cancer.gov/Lasp
