Supplemental Data. Kobayashi et al. (2012). Plant Cell 10.1105/tpc.111.092254
Supplemental Figure 1. Chlorophyll content in shoots of auxin signaling mutants.
Chlorophyll contents in shoots of 14-day-old wild-type Col and auxin signaling mutants,
msg2-1, slr-1, and arf7 arf19, were determined as described in Methods. There were no
significant differences between Col and these mutants. Values are the means ± SD (n >
3).
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Supplemental Data. Kobayashi et al. (2012). Plant Cell 10.1105/tpc.111.092254
Supplemental Figure 2. Chlorophyll accumulation in the stele of the primary root.
(A) Fluorescence from GFP (green) and chlorophyll (red) was detected in the primary
root of 14-day-old SCRpro:GFP seedlings at 1.0~6.0 cm from the root-hypocotyl
junction by a confocal laser scanning microscope (LSM700; Carl Zeiss). The laser
intensity and detection gain of microscopes was fixed during a series of the analyses to
compare fluorescence intensity among different samples. (B) Images for both GFP and
chlorophyll fluorescence in SCRpro:GFP roots were maximally enhanced by a software
program (ZEN 2009; Carl Zeiss) and merged with differential interference contrast
images (Merged image). Bars = 100 μm.
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Supplemental Data. Kobayashi et al. (2012). Plant Cell 10.1105/tpc.111.092254
Supplemental Figure 3. Promoter activities of IAA14, SHR, and SCR in the mature
root.
Promoter activities of IAA14, SHR, and SCR were observed in roots of 8-day-old and
14-day-old seedlings. Promoter activities of IAA14 and SHR were detected
histochemically by GUS staining in transgenic lines carrying the IAA14pro:GUS (Fukaki
et al., 2002) and SHRpro:GUS constructs (Helariutta et al., 2000), respectively, while
those of SCR were detected in a GUS enhancer trap line (End199) in which GUS
activity represents SCR expression (Di Laurenzio et al., 1996; Malamy and Benfey,
1997). An inset shows GUS staining in the mature primary root region of the 14-day-old
IAA14pro:GUS seedling. Bars = 1.0 mm (0.1 mm for the inset). GUS staining was
performed as described (Fukaki et al., 2002).
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Supplemental Data. Kobayashi et al. (2012). Plant Cell 10.1105/tpc.111.092254
Supplemental Figure 4. Chlorophyll autofluorescence in roots expressing the stabilized
mutant IAA14 protein.
Mutant IAA14 protein (mIAA14) was expressed in 21-day-old seedlings via the
indicated promoter in the absence (–) or presence (+) of 1 μM dexamethasone (DEX).
Intact roots at approximately 2.0 cm from the root-hypocotyl junction of each transgenic
plant were examined using a confocal laser scanning microscope (LSM 510; Carl Zeiss).
Merged images represent fluorescent micrographs merged with optical images. Bars =
200 μm.
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Supplemental Data. Kobayashi et al. (2012). Plant Cell 10.1105/tpc.111.092254
Supplemental Figure 5. Spatial images of maximum quantum yield (Fv/Fm) in roots.
Twenty-one-day-old seedlings of wild-type Col, slr-1, ahk2 ahk3, and hy5-215 were
used for the assay. Wild-type seedlings were grown for 7 days in the absence (Col) or
presence (Col + BA) of 1 μM 6-benzyladenine before the assay. Detached roots on agar
plates were placed in a fluorescence imaging system (FluorCam 800MF, Photon System
Instruments) and exposed to actinic light for 5 s (Electronic shutter = 8, sensitivity = 25,
and irradiance = 75) following dark adaptation for 5 min. The image of maximum
quantum yield shown in (B) was obtained by monitoring Chl fluorescence using
software (FluoWin 3.6, Photon System Instruments). The figure in (A) shows a merged
image with the image of maximum quantum yield (B) and the photograph of the roots
captured by a digital camera (C).
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Supplemental Data. Kobayashi et al. (2012). Plant Cell 10.1105/tpc.111.092254
Supplemental Figure 6. Plastid distribution in the primary root of wild type and slr-1.
(A) A cross-section of the primary root of 21-day-old wild-type Col at 4 cm from the
root-hypocotyl junction was observed by transmission electron microscopy. Plastid
development was undetectable in outer vacuolated cells. (B) Plastids with thylakoid
membranes are indicated by arrowheads in the Col root shown in (A). (C) Plastid
distribution in the primary root of 21-day-old slr-1 was observed at the same region as
that in the Col root in (B). Bars =10 μm.
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Supplemental Data. Kobayashi et al. (2012). Plant Cell 10.1105/tpc.111.092254
Supplemental Table 1. Oligonucleotide primers used in this study
Gene
Forward Primer (5'→3')
Reverse Primer (5'→3')
Use
ACTIN8
ACTGTGCCTATCTACGAGGGTTTC
CCCGTTCTGCTGTTGTGGT
qRT-PCR
HEMA1
TAAGATTAGCTTCCCCACAAACTC
AGCTCGCTTATAGCTTCACAACAC
qRT-PCR
CHLH
TGGTAGAGAGACAGAAGCTCGAAA
CCAAAGAACCTGCCCAAGAG
qRT-PCR
CHL27
TCAAGACCGATTACAACCAGACA
CGCTCAAGGAACTCAACGAAG
qRT-PCR
CHLP
ACCAGAAACAGAGCTAAGGACAAGA
GCATCACCTACAAGAGCCACAC
qRT-PCR
GLK1
AAGGGAAGAGAAAGGTGAAGGTG
GTTGTGACGAGTGAGACAATGGA
qRT-PCR
GLK2
TAGCGGAAGAGATGAGGAACAAG
TAAAACCACCGTCACCACCA
qRT-PCR
LHCA4
AACCCGCTTAACTTTGCTCCTAC
CAAACCCTAAGAATGCCAACATC
qRT-PCR
LHCB4
CCACTCCACACCACCATCA
CAAACAAGCAAAAGCCCAAAG
qRT-PCR
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