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Samenvatting Cell and Gene Therapy
- Établissement
- Universiteit Gent (UGent)
Zeer uitgebreide samenvatting die alle slides en zéér uitgebreide notities van de lessen bevat. Behaald resultaat 17/20.
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PART 1+2: MOUSE MODELSMus musculus
Introduction
General
– One of the most important model organisms from a biomedical perspective (mammal!)
– Many advantages compared to other mammals:
o Small adult body size
o Easy to maintain and to reproduce
o Short life cycle (2 months)
o One nest per month (10-15 young)
o Appropriate for genetic research (transgenic mice, mutant strains as a model for
human diseases)
– Disadvantage: price
Strains
– Mice have been inbred to produce hundreds of strains that are homozygous at all loci
– An inbred strain is designated by a unique brief symbol made up from uppercase, Roman,
letters or a combination of letters and numbers beginning with a letter
– Examples: A, C57BL/6, BALB/c, CH3…
Well defined mice
– Inbred strain: A strain that is essentially homozygous at all loci
– In mice, it requires brother-sister matings for at least 20 sequential generations
1
, – Within an inbred strain, individuals are
genetically identical to one another " through
different inbred strains, each carry a unique set
of alleles
– To be sure to have well defined mice, buy them
from a specific company " Every 5 or 10
passages, a new embryo from the (frozen) stock
is taken in this company
General anatomy
" see link in slides for the anatomy
" E.g. E11.5 = embryo is 11.5 days old
Reproduction
Characteristics of reproduction in laboratory mice:
– General similarity to those found in many other mammals
– Polyestrous (having several estrus cycles annually or during a breeding season)
– Spontaneous ovulators – estrous cycle is 4 days (but is influenced by environmental factors
such as light) " in human 28 days!
– Sexual maturity = 5-8 weeks " then they are sexually active
– Formation of luteal tissue is induced by mating
– Like many other rodents: high reproductive potential
– Most aspects of mouse reproduction are amenable to alteration by one or many
environmental factors (light, temperature, presence/absence of other animals…)
– Important source of variability resides in interaction between genetic background and
environment: strains differ in their reproductive responses to the environment
– Reproduction: viviparous (no eggs, living yougs) " the youngs are red and when they start to
drink milk, their belly will turn white " this is a sign that they are healthy
2
, – Gestation = 20 days
– Number of offspring = 2-15
Sperm production
– Diploid spermatogonial stem cells – lifetime supply
– 12 stages of spermatogenesis in the seminiferous tubules " 35
days to complete the development
– Transport to epididymis for storage and maturation (motility)
– For IVF, collect sperm from the epididymis
Oocyte (Egg) production
– Females are born with finite numbers of oocytes
– At 6 weeks, each ovary has 10.000 oocytes
– 6-16 oocytes ovulate over 2-3 hours every 4-5 days in mice
The estrus cycle
– Important to know in which stage of
the cycle the female is
– Whole cycle is 4 days
– To get females pregnant, they should
be in the pro-estrus or estrus-
ovulation phase
– The different phases of the estrous
cycle are visible on the outside vagina
of the mouse
– To get a female pregnant (timed mating):
o Select females that are in pro-estrus
o Put them together with the male (during the night)
o Next day you check if there is a ‘plug’ in the vagina of the female (asap because plug
disappears)
o If you see the plug you know that the female is possibly pregnant " plug does NOT
guarantee pregnancy, f there is no plug, it is still possible that the mouse is pregnant
o From day 10, the female is getting thicker (put it on a balance)
3
, – The pregnancy starts at night, at noon the embryos are 0.5 day old " If you want to inject
something, you know exactly when to do this
example calculation/use
" During the night, the
pregnancy starts – at 12 the
female is half a day
pregnant " embryo is 0,5
days old
" SO: if you want to inject
the mice " timing needs to
be exact " every day at
noon = embryo gets an
injection every day exactly
" Look at female!
Put it on balance from day 8
on " increase in weight
Mating options
– Pair: one feale x one male
– Trio: two females x one male (same cage)
– Harem: single male x more female " not recommended
– Male rotation: two females x male (week 1), same male, two new females (week 2)
o Single mutant male, need many offspring
o Male has a very short lifespan
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