Mouse Reproduction

Reproductive Data for Mice

Sexual maturity
Estrous cycle
Gestation
Weaning age
dependant)
Reproductive life
Life span

40-60 days
4 - 5 days
21 days
21-30 days (this is strain
+ or - 8 months
1 - 3 years

(4-6 litters)

Pups begin eating dry food at about 12 -14 days but

are still largely dependant on their mothers milk for the
bulk of their liquid and nutritional requirements.

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`

Timed mating

Observed for presence

of vaginal plugs
Monogamous pairs 1 male to 1 female
Trios
1 male to 2
females
Harem Mating
1 male to 3+
females
If a harem mating system is used, it is VERY
important to REMOVE pregnant females from the
group cage prior to their delivery.

It is important to remember that when removing

a male from the breeding cage, he should be
housed separately from other males. Males that
have been used for breeding will become very
aggressive and fight with other males sometime
injuring each other severely if returned to a
group environment. Some strains of females
can become aggressive as well, but this is the
exception.
The females have a tendency to be more
compatible than males when returned to a
group environment.

The drawing
at the top
illustrates the
difference in
male and
female pups.
The drawing
at the bottom
illustrates the
difference in
adults.

Male

Female
`

In the adult
mouse, male and
female are easily
differentiated.
The mouse on
the left is male.
The mouse on
the right is
female.

Male
Female

With some
degree of
experience,
very young
mice can be
sexed by
comparing
them side by
side and
noting the
difference in
the distance in
the anogenital
area.

Female

Male

The difference
becomes
more
apparent after
a few days of
age. Darker
mice are more
difficult to
differentiate
than light
colored mice.

Both of these mice

are in estrus they
are in different
stages.
The mouse on the
top would be
chosen for an
immediate breeding
pair set up.
The mouse on the
bottom is in an
earlier stage of
estrus.

Estrus
Note the swollen, moist
and open appearance to the
vaginal opening; coloration
of mucus membranes is pale.

Non-Estrus
Note the closed vaginal
opening and pale mucus
membrane color.

Again, note
the swollen
moist
appearance of
the tissues
surrounding
the vaginal
opening,
bright pink to
purple in
coloration.

Different Stages:

Subtle differences
are evident between
these mice.
Both mice would be
suitable for pairing
with a male if you are
setting up timedpregnancies.
However, the
mouse on the Right
has a higher
probability of
plugging.

Left

Right

Swollen vaginal tissue

Gaping vaginal opening
Moist tissue
Bright pink or purplish
mucous membranes

Closed

vaginal opening
Dry tissues
No swelling evident
Pale mucus membranes

The copulatory plug (most often referred

to as the vaginal plug), is made of several
different components---sperm, ejaculate
fluids, and mucus are primary substances
that harden within the cervix.
The plug forms following copulation and is
visible within, or slightly protruding from,
the vaginal opening. Its presence indicates
a successful mating but does not
guarantee that conception has occurred.

Vaginal plugs are

NOT a guarantee of
pregnancy. They are
only an indication
that copulation has
taken place.
The best time to look
for plugs is in the
morning since mating
usually occurs during
the dark cycle and
plugs only remain in
place for 8 -12 hours

Vaginal plugs can be very

easy to find, or they
can be very difficult. It
depends on the strain
of mouse and the
length of time that has
elapsed since the plug
was been formed.
Some strains do not
produce as much
seminal fluid as others
making the plugs more
difficult to detect.

The plug is opaque,

whitish, and has a waxy
consistency. It can vary
in color from snow
white to yellow. It is
generally easy to
identify while still
within the vaginal
opening. It may also be
identified in the
bedding as a large waxy
chunk, if one checks
closely. Generally, plugs
are expelled close to
the nest/sleep area.

Sometimes the plug can be

difficult to visualize.
To aid with its identification,
gently palpate the area to
help distend and partially
expel the plug.
The use of forceps is not
recommended. If forceps
are used, extreme care
must be taken and the plug
should never be forcibly
removed.

Once a plug has been detected, the female

should be removed from the male and placed
in her own cage to ensure that the potential
gestation period is as accurate as possible.
Leaving her exposed to the male after the
observation of the vaginal plug can result in
subsequent breeding and inaccuracy of the
expected birthing dates.
Remember that the vaginal plug is only a sign
that mating has occurred, it is NOT a
guarantee of pregnancy.

After she has

given birth, the
mother will
remain on the
nest to care for
her young.
Females that are
first-time
mothers are
sometimes very
careless with
their young and
mortality can be
very high.

First-time mothers are sometimes very

careless with their young and mortality can
be very high.
It is not unusual for a first time mother to
cannibalize her litter. In many cases, future
litters are successful. However, if the female
continues to be a poor mother, it is best if
she can be removed from the breeding
program.
Females with a strong maternal instinct seem
to pass this trait on to their offspring. The
opposite tends to be true as well.

It should be noted at this time that breeding some

transgenic lines can present unique challenges.
There can be fertility problems with both males and
females that require you to use a heterozygote
breeding system rather than homozygote.
Some females will be able to carry to term but not
be able to produce milk for their young.
Some genes may be embryonic lethal and while the
female becomes pregnant, she will abort or
reabsorb the pups prior to birth.
Each line is different, so research into the potential
problems is important.

Newborn, or 1
day old mice
are very red,
helpless and
hairless. The
ears and eyes
are shut. If
they are
nursing, milk
can easily be
seen in the
stomach.

At day 2 the
pups are
less red,
more pink
Milk can still
be seen in
the stomach

By day 3, the
ear flaps will
begin to open
If the pups
are going to
be black or
agouti, the
skin pigment
begins to
develop

On day 4 the
ears are erect
and milk is
no longer
visible in the
stomach

By day 5, the
pups will
begin to
develop light,
fuzzy dorsal
fur

At day 6 the
fur is thicker
across the
shoulders
and the coat
color may be
evident

By day 7
the back of
the pups is
completely
covered in
fur.

Day 8, the
belly is
beginning
to show
fur

`
`

On day 9 the
fur is thicker.
Females will
have five
pairs of
conspicuous
nipples while
the males will
be much less
obvious

Day 10, the

fur growth is
complete and
muscular
activity is
increased

By day 11 the
teeth are
beginning to
erupt and the
eyes start to
open.

At days 12,13
and 14, the
eyes are
mostly open
and they begin
to nibble of
solid food
Even though
they may be
nibbling at
solid food they
are still almost
totally
dependant on
their mother.

These pups are 3 weeks of

age and ready to be
weaned. When weaned, you
should place food on the
bottom of the cage. If they
are on automated water,
you should place a water
bottle on the cage for a few
days as an alternate source
of water in case they are
slow to catch on to the lixit
as the primary water
source. Always keep a close
eye on newly weaned mice.

This is a
comparison
of a four
week old
mouse and
an adult
mouse.
Note the
difference
in the
shape of
the head.

It is extremely important that you provide food and a

water source to newly weaned pups to help ensure their
survival.
Food must be placed in the bottom of the cage or
dough diet provided on cage floor.
A WATER BOTTLE must be placed on the cage, including
those cages hooked up to automated watering, or gel
pack placed on the floor of the cage. Using an
automated watering source is not second nature to
weanlings, they must LEARN to use it!
DO NOT UNDER ESTIMATE THE IMPORTANCE OF THESE
ACTIONS. It could mean life or death for your
weanlings.

The best way to place regular rodent chow in the bottom

of weanling cages is to moisten it with water from a
clean water bottle NOT TAP WATER. While many
weanling mice are able to gnaw on dry rodent chow on
the cage floor, moistening the food increases their
ability to eat it thereby increasing the chance for
survival.
Place a small amount of pellets (5-10 depending on the
number of weanlings) in a dish and pour a small amount
of water from a clean water bottle over them.
You want the food moistened and somewhat soft but not
soaked. Too much water will cause the food to spoil.
You will have to repeat this process for several days until
the mice grow stronger and be able to eat the food
without the extra moisture.

Large pellets should be

broken in half and
placed in a small dish
or container. Do not
just drop water on the
food in the cage
bottom since the
bedding will absorb the
water before the food
does.

`
`

Place small pieces of dry food into a small dish.

Add about inch or so of water (depending on the number of
pellets) from a clean water bottle, autoclaved water or RO
water is fine.
DO NOT USE TAP WATER or a water bottle that has been used
by other animals.
Allow water to be absorbed by food, this takes about 3-5
minutes or so.
Moistened food can now be placed in the bottom of the
weanling cage.
Dry food

Moistened food,
Note darker
color and fuller
shape

Food and Nestlet

on cage floor

Dough Diet on
cage floor

Moistened food pellets

Dough Diet and gel
pack on cage floor

Water bottle
within reach of
weanlings, food
and Nestlet on
cage floor

The reproductive life of any particular

mouse strain will vary. There are many
factors that go into the equation. Litters
from Transgenic mouse strains can vary
from as few as one or two to as many as
six to eight. Carefully kept breeding
records are essential in the success of any
colony.

Replacement of non-productive animals,

good record keeping, including production
indices, timely weaning of litters and close
observation will aid in the overall success
of the colony. Remember to enter your
weaning dates and the other data in the
weaning book. Contact the main DLAR
office for additional weaning cards if you
need them.
PLEASE DO NOT REUSE CAGECARDS.

The amount of floor space for a mouse is

determined by The Guide for the Care and
Use of Laboratory Animals published by The
National Research Council
Guidelines for cage populations under
different circumstances are outlined by the
institution based on these guidelines.

No more than 4 adult mice or weanlings in

a static Micro-isolator or small PIV cage
(144 cages to a rack) or no more than 5
adult mice or weanlings in a large PIV cage
(112 cages to a rack or the green line) are
to be housed in the same cage.

`
`

females

Cages with two (2)

and 13 or
more pups
Cages with two or more females and two or
more litters of different ages.
Of Special Note: Cross fostering of pups, in
most cases, results in litters of different ages
in the same cage. These cages must be
clearly labeled indicating that cross fostering
is occurring. Be sure to check to see if IACUC
approval is needed BEFORE you cross foster.

Cages with a singly-housed female

and her litter when the pups are 29
days of age or older and/or
immediately upon the birth of a
second litter into the cage.

2 adult mice and 1 (one) litter

when the pups are 22 days of age or older
Cages with

must be weaned
The older pups must be weaned immediately
upon the birth of a second litter into the
cage (usually when older litter is in excess of
19 days of age if adults are a breeding pair).

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Dr. Jeanie Kincer

Dr. Jeff Smiley
Peggy Hankes-Pea
Glenn Florence
April Davis
JoeAnn Croxford

Office
323-5469
323-0289
323-6015
257-1026
323-1547
323-3616

Pager
859-330-2082
859-330-2099
859-330-1571
859-330-1504
859-330-0388

Maintaining a rodent breeding colony requires

a great deal of time and commitment. Will you
be able to devote the time and energy it takes
to maintain the colony efficiently?
Do you really need to establish the colony
yourself or are the animals available from
another source, often saving valuable time and
resources.
Remember that older animals, over 6-8 months
of age, do not make good breeding prospects
and will often delay your study because of their
failure to reproduce.
Over handling and disturbing the female
shortly before she gives birth, or after she has
had a litter, may increase the risk of problems
developing.

Occasionally, for one reason or another, you

will run into a problem where the mothers of
the litters are unable to properly care for their
young. Do you have an alternate means to
care for them in mind?
Fostering can be a fairly simple task, but you
will need to plan ahead for these times.
While not totally successful 100 percent of the
time, fostering mice can be relatively
successful if a few basic practices are
followed.

 The foster mom must have a healthy, well fed litter of her own.
The litter to be fostered should be within one or two days of
the age of the foster moms own litter.
If attempting to cross foster, make sure these cages are clearly
marked and check to see if IACUC approval is needed.
It helps if the pups are a different color then the litter to be
fostered.
The foster litter should be no larger than the natural litter, and
here is where it gets a little tricky. If you have a large litter of
pups that need to be fostered, you should split them between
several foster moms. You need to be careful to record all of
the information including the color and sex as well as the
original genetic information about the foster litter when you
place them with the foster mom.

If the dam dies or cannot or will not feed her new

pups, here are some steps you can take to try to
save the litter. Unfortunately, not all attempts to
foster a litter will be successful.
Once a foster mom is identified, she should be
removed from her home cage and placed in a new
cage.
If you will be placing the fosterlings with her
existing litter, you should put them in the nesting
material with their foster siblings and cover them
with some of the soiled bedding to transfer the
foster litter and moms scent to them.
Do not place the joined litter and foster mom in a
clean cage. This greatly increases the chances of
total rejection.

When returning the foster mom to the newly

formed litter, you can allow her to urinate on the
pups by holding her over them for a moment or
two, or if she has urinated in the holding cage
use a cotton swab to recover some of the scent
and place it on her nose. This is not a
requirement.
Some strains of mice make better foster moms
than others. Many transgenic strains are not
good candidates for accepting fostering litters.
Some transgenic strains have their own rearing
difficulties. Balb/c, FVB, and Swiss Websters
are excellent foster moms.

Not all foster moms will accept the fosterlings

The foster pups are too cold or lethargic and do not
respond well to stimulation. They should be warm and
wiggly. The foster mom will reject pups that are cold to
the touch, are obviously weak and are unable to suckle.
She will be very confused and may reject pups that are
either much larger or much smaller than her own litter,
even if the natural litter is removed.
The birth cage or nursing cage for her own litter has been
changed and there is not enough scent from her own
litter to mask the foster pups odor.
The foster mom is a first time mom herself.

 You should monitor the newly formed litters

very carefully and very often to be certain the
pups are being accepted and are being fed.
This is usually evident within the first hour or
two.

Of Great Importance- If the foster mom is

not taking care of the fosterling, do not just

let them fade away. They should be humanely
euthanized if it is obvious they are not being
fed and cared for by the foster mom.

There are many factors that determine how many

animals you really need, and they should all be
included in your breeding plan and the number
of animals listed in your IACUC approved
protocol. You should, also, include the space
requirements and costs associated with
maintaining them.
Please remember that when you are managing a
breeding colony, your goal is to meet the needs
of your study as it relates to your protocol.

Remember

^ Be Nice to the Mice ]