Definition: Transgenic Animal

1
Definition Transgenic Animal

• Animal in which a segment of DNA has been
  physically inserted into the genome.
• The genome of all cells of the organism contains
  the DNA segment.
• The DNA segment is present in the germ line so it
  can be transmitted to progeny as a Mendelian
  trait.

2
Retroviral Mediated Gene Transfer
Mouse embryos
MoMuLV Producing Cells
Insertion of viral DNA containing transgene into
embryos
Transgenic Mouse
3
PMS HCG to superovulate
1-cell pronuclear stage mouse embryo.
X
2-cell stage embryo
Injection Pipette
Holding Pipette
Oviduct Transfer


? Pregnant Foster Mother
4
Events During Transgenesis
E
1
E
2
E
2
1
D
A
Y
1
2
3
4
5
B
i
r
t
h
E
m
b
r
y
o
R
e
c
o
v
e
r
y
hCG
PMS
1
2

A
M
1
2

A
M
1
2

A
M
1
2

A
M
1
2

A
M
1
2

A
M
1
2

A
M
5
Embryonic Stem Cells

• Pluripotent stem cells isolated form the inner
  cell mass of a blastocyst stage mouse embryo.
• Cells retain ability to differentiate into all
  organs and cells.
• Cells can be propagated under conditions in vitro
  that allow them to retain their totipotency-
  prevent differentiation in vitro.
• Can be induced to differentiate in vitro into a
  number of cell types. ES cells can mimic early
  mammalian development in vitro.
• Can be manipulated in vitro to alter the genome
  through homologous recombination.

6
ES cell Culture
Targeted ES Cells
Targeted ES Cells Injected into Blastocyst
Uterine Transfer
Chimera
? Pregnant Foster Mother
Blastocyst derived from mouse with white coat.
X
Germline transmission of ES cell-derived genome.
Breed to WT white mouse
7
Gene Knockout by Homologous RecombinationUsing
an O or Insertion Type Construct
A B C D E F G H I
X
neo
A B C D
E F G H I
neo
8
Gene Knockout by Homologous RecombinationUsing
an W or Replacement Type Construct
A B C D E F G H I
X
X
TK
neo
A B
G H I
neo