Cloning Flashcards
(48 cards)
Clone
an organism descended asexually from a single ancestor.
-sending the cells back.
SCNT
the transfer of a nucleus from a somatic cells into an oocyte from which the nucleus has been removed.
Reproductive cloning
the genetic duplication of an existing organism.
Therapeutic cloning
production of autologous undifferentiated cells for therapeutic uses.
Natural cloning
• identical (monozygotic) twins (Embryo expands and collapses and breaks (during the blastocyst stage) or during hatching)
• polyembryony (two or more embryos
developing from a single fertilized egg) (ie the zygote splits)
Artificial cloning
SCNT,
- embryo splitting
- nuclear transfer
polyembryony
more than one embryo from the same zygote
How many cells need to be present in the blasocyst to clone?
No more than 4.
-The cells become too small to separate and develop because they don’t have enough cytoplasm.
x
can clone from any stage. adult cells are more differentiated than embryonic cells.
embryo splitting
if you take a good quality embryo and cut it in 2, 50% of the time you will producing a calf
ET has a 60% of producing a calf.
-cut at the morula stage (dont need to worry about cutting the wrong place because the cell is not very differentiated)(ie having too much fetus not enough placenta etc).
-worry at the blastocyst stage
x
oocytes need to be competent - to support embryo development and cell reprogramming
- no sperm is involved now : sperm induces the release of Ca to degrade MPF is not there, so this won’t happen.
- culture the nuclear donor cells in vitro
x
- mature the oocyte
2. remove the chromatin via aspiration
problems of SCNT
- Lower embryo development
- Lower pregnancy rate after embryo transfer
- Higher fetal mortality
- Lower viability of cloned animals
- Despite of the low efficiency, most cloned animals can grow, reproduce and produce normal offspring
However, the most of the ones that are born are normal and the ones that are not normal will produce normal offspring
probability of SCNT
less than 10%
Problems of SCNT cloned animals at the perinatal stage ie the ones that we have chosen to develop.
• Malformations • Large (in ruminants) or small (swine) offspring syndrome • Respiratory insufficiency • Hepatic congestion / insufficiency • Immunodeficiency • Multisystemic dysfunctions more than 60% are capable to develop
Effect of abnormal cell cycle interactions on chromatin integrity in SCNT cloned embryos
fragmentation
condensation
multiple nuclei
which stage is the easiest to synchronize?
G1
What happens when you use metaphase 2 cells as donors?
MPF levels are high, so the chromatin will condense. then depending on the donor nucleus the ploidy may be abnormal if they are replicated right away.
what happens when you use interphase cells as donors?
MPF levels are low. the interaction between the cytoplasm and the nucleus is decreased. No abnormal ploidy because the chromatin did not condense
what do we want to maintain when we fuse for SCNT?
- potential for reprogramming
- chromatin integrity
- embryonic ploidy
Effects of cell cycle interactions between nuclear donor cells and host oocytes
- Effects on chromatin integrity - incompatibilities between the cell cycle stage of nuclear donor cells and host oocytes may cause DNA damage
- Effects on nuclear reprogramming – the cell cycle stage of host oocytes and donor cells affect nuclear reprogramming in SCNT embryos
- Effects on DNA replication - cell cycle incompatibilities may cause DNA re-replication.
trend between NT efficiency and differentiation of donor cells?
as cells become more differentiated Nuclear Transfer efficiency decreases
What is the cause of the abnormal gene expression in cloned embryos ?
Epigenetics
what is the general trend of epigentic markers between more differentiated cells and less differentiated cells?
less differentiation have more Acetylation
more differentiation have more Methylation. The Methylation moves down the chain (attached to a lysine further down)
so to reprogram it we must remove CH4 and ad Ac
