whether
interaction
between
co
cop1
also
occurred
vivo
plant
cells
tested
using
fluorescent
resonance
energy
transfer
fret
microprojectile
bombardment
used
co-express
cyan
fluorescent
protein
cfp
cop1
yellow
fluorescent
protein
yfp
co
leaf
epidermal
cells
arabidopsis
cfp
cop1
yfp
co
colocalized
nucleus
also
colocalized
speckles
within
nucleus
figure
4a
physical
interaction
cfp
cop1
yfp
co
tested
measuring
fret
using
photoacceptor
bleaching
previously
described
wenkel
et
al
figure
4c
quantification
fret
signals
demonstrated
fret
occurred
between
yfp
co
cfp
cop1
both
nucleus
specifically
nuclear
speckles
figure
4c
control
experiments
using
yfp
cfp
yfp
co
cfp
yfp
cfp
cop1
fret
detected
significantly
lower
levels
figure
4c
experiments
demonstrate
yfp
co
cfp
cop1
colocalize
physically
interact
nuclei
plant
cells
cop1
full-length
cdna
isolated
rt
pcr
produced
entry
clone
bp
reaction
gateway
system
from
invitrogen
entry
clone
utilized
construction
destination
vectors
plant
transformation
fret
experiments
vitro-binding
assay
plasmids
plant
transformation
introduced
agrobacterium
strain
gv3101
pmp90rk
transformed
wt
columbia
cop1
suc2
co
et
al
plants
floral
dip
method
clough
bent
quantification
fret
vivo
between
cfp
co
yfp
cop1
yfp
co
detected
increase
cfp
fluorescence
photobleaching
yfp
quantification
fret
efficiencies
acceptor
photobleaching
measured
nuclei
nuclear
speckles
data
mean
s.d.
cells
from
three
separate
experiments
visualization
increase
cfp
fluorescence
yfp
photobleaching
left-hand
panel
cells
expressing
cfp
cop1
yfp
exerts
effect
negative
control
right-hand
panel
cells
expressing
cfp
cop1
yfp
co.
scale
barfluorescence
resonance
energy
transfer
fret
pairs
gfp/dsred
analysed
using
confocal
laser-scanning
microscope
zeiss
lsm510
meta
fret
measurements
dsred
emission
zero
contribution
from
gfp
accomplished
described
erickson
et
al.
using
following
settings
excitation
nm
emission
filters
bp
nm
gfp
bp
nm
dsred
fret
detection
tobacco
leaf
epidermal
cells
co-expressing
gfp
atebp
acbp4
dsred
differential
interference
contrast
image
d-f
green
channel
shows
gfp
atebp
red
channel
shows
fret
signal
acbp4
dsred
co-localization
two
signals
indicated
yellow
colour
merged
images
fret
analysis
cells
co-expressing
gfp
atebp
acbp4
dsred
only
gfp
atebp
green
fluorescence
fig.
2d
also
acbp4
dsred
red
fluorescence
fig.
2e
overlapped
gfp
signals
fig.
2f
detected
indicating
fret
occurred
between
gfp
atebp
acbp4
dsred