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Wavetek 131A Manual do Utilizador Página 4

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Homozygous LGI1
/
mice display
early onset spontaneous seizures
The behaviour and appearance of LGI1
/
mice at birth did not
differ from that of LGI1
+/
and wild-type littermates. In the
second postnatal week, however, both male and female LGI1
/
mice began to exhibit frequent spontaneous seizures. Seizures
were first observed at postnatal day 10, especially during cage
changing and handling. They consisted of a behavioural sequence
including (i) movement arrest, sometimes associated with limb
A
Cre
recombinase
Floxed allele
Knockout allele
TGA
ATG
LoxP
LoxP_FRTNeoLoxP_FRTP
1234567 8
557 amino acids
ATG
Premature TGA
12345 8
179 amino acids
64 kDa
51 kDa
B
DE
97 kDa
64 kDa
51 kDa
51 kDa
39 kDa
28 kDa
19 kDa
α-tubulin
LGI1
51 kDa
39 kDa
28 kDa
19 kDa
α-tubulin
α-tubulin
LGI1
LGI1
LGI1+/+
LGI1+/-
LGI1-/-
LGI1+/+
LGI1+/-
LGI1-/-
C
97 kDa
E
1
6
P
0
P
3
P
6
P
9
P
1
2
P
1
6
A
d
u
l
t
51 kDa
64 kDa
B
r
a
i
n
Sp
i
n
a
l
c
o
r
d
M
u
s
c
l
e
H
ea
r
t
L
u
n
g
S
t
o
m
a
c
h
S
p
l
e
e
n
K
id
n
e
y
L
i
v
e
r
LGI1
Ponceau S staining
64 kDa
Figure 1 Generation of the LGI1 knockout mouse. (A) Diagram showing LoxP and LoxP-Neo-LoxP sequences indicated by triangles
(floxed allele). Exons 6 and 7 were removed by crossing the LGI1
loxP
mice with a PGK-Cre strain (knockout allele). (B–C) Western blots
demonstrating the absence of native LGI1 protein in whole brain lysate of LGI1
/
mouse (at postnatal day 13) using the ab30868
antibody (B), and spinal cord lysate of LGI1
/
mouse using the sc-28238 antibody (C). Equal amounts of total protein were loaded as
demonstrated by -tubulin levels. (D) Western blot showing LGI1 expression during embryonic and postnatal development on C57BL/6
mouse brain lysate (ab30868) and -tubulin used for internal control. (E) Western blot showing restricted expression of LGI1 in the brain
and spinal cord of a postnatal day 15 wild-type mouse (ab30868). Equal amounts of total protein were loaded and Ponceau S staining was
used as internal loading control.
2752 | Brain 2010: 133; 2749–2762 E. Chabrol et al.
by guest on May 15, 2015Downloaded from
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