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Tips for Successful Breeding and Colony Management. Office of Lab Animal Resources University of Colorado Denver AMC Tracy Haney, CVT, RLAT Holly Goold, CVT, RALAT. Environmental Factors. Drop in production (October-March) Even though mice are in a controlled environment

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Tips for successful breeding and colony management

Tips for Successful Breeding and Colony Management

Office of Lab Animal Resources

University of Colorado Denver AMC

Tracy Haney, CVT, RLAT

Holly Goold, CVT, RALAT


Environmental factors
Environmental Factors

  • Drop in production (October-March)

    • Even though mice are in a controlled environment

      • Light cycles, temperature, humidity, etc

    • Smaller, less frequent litters.


External factors
External factors

  • Odor from staff (perfume/cologne, cigarette smoke)

  • Entering housing room after hours (8p-6a)

    • Mice mate 5-6 hours after dark cycle starts

  • Conversations/music/phones in housing room

  • Amount of traffic in housing room

  • Shoving cages anywhere on rack

  • Inexperienced /unconfident handling

    of mice


Cage location
Cage location

  • In room

    • If possible, away from door and ATS

  • On rack

    • Lowest row on rack, away from lights

    • Keep separate from experimental mice

      • Less cage movement


Enrichment
Enrichment

  • Helps decrease stress/increase pup survival

  • Manage their environment

  • Mimics natural nest

    • Brown paper shred

    • Mouse huts/paper towel rolls

  • Con

    • Remove before cage wash if combined with nestlet

    • Must use ~6-8 grams of shred

    • Hard to visualize litters


Breeder set up
Breeder Set-up

  • Setting Breeders

    • Mice and Rats

      • Females: 5 weeks to 8-10 months

        • Breed females before 3 months of age

          • better fertility

        • Litter sizes decrease with age

        • Genetic issues

          • Poor lactation in homozygous moms

      • Males: 6 weeks to 12-18 months

      • Set-up

        • Set in a clean cage

        • Add enrichment/breeder chow

        • “Charged” cage


Types of matings
Types of Matings

  • Continuous pair

    • Pros

      • Post-partum estrous cycle

      • Litters every 21 days

    • Cons

      • Possibility of two litters in the cage

  • Non-Contiuous

    • Pros

      • No double litters

    • Con

      • Have to remove the male

      • Less frequent litters

  • Triad

    • Pros

      • Sister females, synced estrous cycle

    • Cons

      • Unrelated females

        • 1 Dominant female

      • Requires protocol approval

      • Male rotated between cages

      • Miss females’ post-partum estrous


Breeding tips
Breeding Tips

  • Lithgow “Check for Pups/Wean” cards are provided in housing rooms

    • Track births, pups lost, and successful weaning

    • Tracking fertility


Breeding tips1
Breeding Tips

  • DO NOT retire old breeding pairs until after confirming successful breeding of new pairs

    • Retiring old breeders

      • Set new breeding pair one month before retiring old

      • Ensure successful breeding before ending old pair

    • Stagger setup

      • Insurance to always have mice


Maintaining a litter
Maintaining a Litter

  • Avoid extra handling of litter

    • < 3 days old

  • Reduce litter size when able

    • One sex vs phenotype

    • Genotype early: 7-10 days

      • Footpad tattoos/tail snips

  • Cage changing

    • Anticipating pup birth

    • Transfer half of the old nest with pups

  • Fostering

  • Keep environmental noise levels “even”


Troubleshooting
Troubleshooting

  • Pup death

    • Stress to female

      • Disturbing the cage frequently

      • Removing male right before or after pup birth

      • Separating females from cage right before or after pup birth

      • Manipulating pups- touching pups within a week of birth

    • First-time mom (C57 background)

    • Transgenic lines

  • *Please remember that you must remain compliant with the IACUC rodent overcrowding policy. If multiple litters are in the cage, you are obligated to separate them for animal welfare reasons regardless of the risk of cannibalism or maternal neglect. Therefore, it is essential that you separate mothers prior to the litter being born.


Breeding specialists
Breeding Specialists

For information on breeding services or training/help needed, please contact:

Tracy Haney for Barrier and R2:

[email protected], 303-724-3982

Holly Goold for RC-1:

[email protected], 303-724-2237



Why do my mice eat their babies diagnosing lactation problems in mice town hall meeting oct 24 2013

Why do my mice eat their babies?!?Diagnosing lactation problems in miceTown Hall meeting, Oct. 24, 2013

Jenifer Monks, Ph.D.

[email protected]


How could my gene mutation be affecting lactation?!?Gene mutations may alter mammary function: off-target

  • Cell cycle, proliferation, cell death-> mammary development

  • Cell polarization, secretion, metabolism-> milk production

  • Neurological/behavioral-> mothering, stress, let-down, feeding behavior in pups

Developmental Stages of the Mammary Gland

Embryonic

Pubertal

Adult

Involution

Pregnancy

Lactation

…use wild-type, heterozygous or hemizygous females whenever possible


Indications the dam may be having trouble feeding her pups
Indications the Dam may be having trouble feeding her pups

  • Normal Estrous

  • Get pregnant, stay pregnant

  • Normal delivery, normal litter size

  • Pups scattered around cage

  • Die peri-partum (shortly after birth) or are cannibalized

  • Failure to thrive

  • Wean runty or with bald butts

  • Only alternate litters survive


Tips for success
Tips for success:

  • Nesting material: thermoregulation, security, pheromones

  • “Do Not Disturb” pink cards peri-partum: reduce stress

http://med.stanford.edu/ism/2012/march/mice.html

Making mice comfy leads to better science, researcher says

“The shape of the nest tells an experienced person whether the animals are too hot or too cold, whether they are sick or whether they are about to give birth,”


Do they have milk bands
Do they have milk bands?

No? dead by day 3



Tips for success1
Tips for success:

3) Careful record keeping

Estrous 4-5 days

Gestation 18-21 days

Lactation 21-28 days

(if housed with male, remove weanling animals at 20 days old)


Gestation time of your strain is known
Gestation time of your strain is known:

A/J

20.5

129

20.2

Balb/c

20.1

C57BL6

19.6

FVB

18.7

Murray SA, Morgan JL, Kane C, Sharma Y, Heffner CS, et al. (2010) Mouse Gestation Length Is Genetically Determined. PLoS ONE 5(8): e12418.


Mouse weighing to track reproduction:Normal gestation is 19 days- Co-housed with male-implantation delay when concurrently pregnant and lactating


Dam litter body weights non invasive monitoring

**

n = 8 litters

Dam-Litter body weights: non-invasive monitoring

mid-pregnancy resorption of litter

Compromised milk production

Lactation Failure and Early weaning by mother


Cross-fostering to improve breeding success

CD-1/ICR & Black Swiss

mice make good foster dams

Timed matings of both strains

Remove transgenic pups from biological mother and give to foster dam as soon after birth as possible

Warm in hand if cold and scattered

Roll in soiled bedding of foster dam

Remove unneeded pups to normalize litter size-10 teats, match natural litter size

Place carefully in nest


Timed mating of mice
Timed mating of mice

Photo courtesy Mouse Fancier website

http://www.fancymicebreeders.com/mousefancierforum/phpBB3/viewtopic.php?f=11&t=756

Four stages of estrous in BALB/cByJ mice. The four stages of estrous are shown for an albino strain (proestrus (A), estrus (B), metestrus (C), diestrus (D)).

Mouse estrous cycle identification tool and images. 2012. PloS one    


Why care about breeding efficiency
Why Care about breeding efficiency?

  • Save time

  • Save money: $0.83/day/cage

  • Better animal care = better science

Tips for Increasing Efficiency and Cutting Costs

  • Do not set up breeders without knowing their genotypes.

  • Know what genotypes you are trying to produce and the number you will need.

    • Overproduction = wasted animals and $$$

    • Underproduction = wasted time

    • Account for seasonal variation and holiday schedules

  • Keep a reserve of breeder-age animals.

  • Retire/replace unproductive breeders (record keeping, 2 consecutive lost litters), unused experimental animals, and animals of useless genotypes.

  • Do not keep singly housed animals

  • Keep a calendar for breeders and experiments

  • * Collaborate *

Know which animals are Raptor Safe

Contact OLAR if you have unused animals that can be donated to training protocols


Still having trouble ask for help
Still having trouble? Ask for help!

  • Online resources-Jackson Laboratories

  • OLAR: Care staff, vet techs, breeding specialists

  • Colleagues

  • …Andrew Lewis

    • Toothless mice

    • Mayonnaise milk

      [email protected]


Website www medschool ucdenver edu transgenics email makeamouse@ucdenver edu

Transgenic and Gene Targeting Core

Website: www.medschool.ucdenver.edu/Transgenics

Email: [email protected]


Transgenic and Gene Targeting Core

www.medschool.ucdenver.edu/Transgenics


Core Personnel

Peter J. Koch, PhD

Professor of Dermatology and Cell & Developmental Biology, Director

The Charles C Gates Center for Regenerative Medicine and Stem Cell Biology

University of Colorado

Saiphone Webb, BS

Senior PRA, Laboratory Manager

Embryo Manipulation Services

Embryonic Stem Cell Development and Production

Abby Zamora, CVT, LAT

PRA

Colony and Data Management

Embryo Manipulation Services

Abhilasha Jain, MS

PRA

Embryonic Stem Cell Development and Production


How You Can Lose Your Mouse Line

  • The transgene stopped working; loss of phenotype (e.g. promoter methylation)

  • There is a change in the phenotype of your mice (e.g. gene drift)

  • A disease outbreak

  • Breeding has stopped and you’re left with only few male and female mice.


Solutions

  • Order replacement mice from a vendor(if the line is available)

  • Request the mice from a colleague working at another University – (this will take time to breed up the mice for experiments)

  • Start over and generate the line from scratch(this will take more time and money)

  • Re-establish mouse line using previous frozen sperm(IVF) or embryos



Transgenic and Gene Targeting Core Rescue Services

www.medschool.ucdenver.edu/Transgenic

[email protected]

Supported by: NIH (SDRC, CCTSI), Gates Center, SOM,

Dermatology Department, Service Fees


?? Questions ?? Rescue Services


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