mounted
cells
visualized
under
fluorescent
microscope
leica
http://www.leica.comsubcellular
localization
endogenous
exogenous
uxt
293t
cells
transfected
top
without
bottom
flag-uxt
immunofluorescentmicroscopy
performed
indicated
primary
antibodies
alternatively
performed
immunofluorescence
analysis
support
speculation
interestingly
transfecting
cells
sirna
against
uxt
stimulating
tnf-α
apparently
decreased
percentage
cells
displayed
focused
nuclear
p65
approximately
control
cells
displayed
p65
inside
nucleus
upon
stimulation
whereas
only
displayed
p65
case
uxt
knockdown
specimen
fig.consistent
data
s329/t331-phosphorylated
mdc1
detected
immunofluorescence
both
control
γ-irradiated
cells
fig
2b
note
absence
staining
cells
treated
mdc1
small
interfering
rna
sirna
confirming
sdtd-phosphorylated
mdc1
present
absence
damage
also
forms
iriffluorescence
microscopy
demonstrating
shift
subcellular
localization
gfp-sed5
from
golgi
control
wt
strains
partially
cytosolic
localization
δget3
strain
both
cytosolic
few
large
puncta
δget1
strains
gfp-sed5
puncta
get
mutants
colocalize
golgi
marker
anp1-rfp
fluorescence
microscopy
demonstrating
colocalization
gfp-sed5
get3-tdrfp
cytosolic
aggregates
form
δget1
background
fluorescence
microscopy
control
wt
δget1
strains
expressing
broad
variety
ta
proteins
gfp-scs2
gfp-sbh1
gfp-ysy6
under
galactose-inducible
gal
promoter
cherry-sbh2
expressed
from
plasmid
under
constitutive
tef2
promoter
fluorescence
microscopy
control
wt
δget1
strains
expressing
two
mitochondrial
ta
proteins
cherry-fis1
cherry-tom22
expressed
from
plasmid
under
constitutive
tef2
promoter
fluorescence
microscopy
showing
localization
gfp-ubc6
mitochondrially
targeted
dsred
mts-rfp
control
wt
δget1
strain
fluorescence
microscopy
time
course
monitoring
subcellular
localizations
peroxisomal
ta
protein
gfp-pex15
well
dsred
targeted
mitochondria
mts-rfp
following
induction
pex15
from
galactose
inducible
promoter
control
wt
get1/2
get3
strainascertain
time
infection
subcellular
compartment
interaction
occurs
asfv
infected
cells
examined
confocal
laser
scanning
microscopy
different
times
post
infection
least
infected
cells
analyzed
each
time
point
hpi
subcellular
distribution
p30
hnrnp-k
analyzed
immunofluorescence
microscopy
asfv
mock
infected
cells
asfv
infection
resulted
intensification
nuclear
cytoplasmic
staining
hnrnp-k
compared
uninfected
cells
nuclei
infected
cells
examined
detail
nuclear
hnrnp-k
detected
characteristic
granular
structures
coincident
areas
completely
devoid
nucleic
acid
staining
interestingly
kind
hnrnp-k
accumulation
also
observed
cells
transiently
expressing
p30
pcmv-p30
mock-infected
cells
suggesting
involvement
p30
changes
fig.
during
asfv
infection
colocalization
p30
hnrnp-k
nucleus
infected
cells
cells
infected
ba71v
strain
analyzed
different
times
post
infection
confocal
immunofluorescence
microscopy
acquiring
optical
sections
from
z-axis
representative
image
infected
cell
hpi
shown
p30
detected
monoclonal
anti-p30
followed
alexa
647-conjugated
goat
anti-mouse
antibody
red
hnrnp-k
anti-hnrnp-k
specific
serum
followed
alexa
488-conjugated
goat
anti-rabbit
antibody
green
discrete
spots
colocalization
both
proteins
orange
discerned
cell
nucleus
asfv
infection
induces
changes
subcellular
distribution
hnrnp-k
vero
cells
either
infected
ba71v
pfu/cell
transfected
pcmv-p30
analyzed
immunofluorescence
microscopy
hpi
post
transfection
respectively
asfv
protein
p30
detected
mouse
anti-p30
monoclonal
antibody
rhodamine
red
conjugated
corresponding
secondary
antibody
red
hnrnp-k
distribution
cell
nucleus
detected
rabbit
antiserum
anti-hnrnp-k
alexa
fluor
conjugated
corresponding
secondary
antibody
green
nuclei
cells
stained
hoechst
accumulation
hnrnp-k
infected
cells
spots
coincident
absence
nucleic
acids
staining
appreciated
ba71v
infected
pcmv-p30
transfected
cells