Genome-Wide Identification, Evolution, and Transcriptional ...
„Genome-Wide Identification and Testing of Superior …€¦ · · 2011-01-03Supplemental...
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Supplemental Material „Genome-Wide Identification and Testing of Superior Reference Genes for Transcript Normalization in Arabidopsis thaliana.“ by Tomasz Czechowski, Mark Stitt, Thomas Altmann, Michael K. Udvardi and Wolf-Rüdiger Scheible
Transcript of „Genome-Wide Identification and Testing of Superior …€¦ · · 2011-01-03Supplemental...
Supplemental Material
„Genome-Wide Identification and Testing of Superior Reference
Genes for Transcript Normalization in Arabidopsis thaliana.“
by Tomasz Czechowski, Mark Stitt, Thomas Altmann, Michael K. Udvardi and
Wolf-Rüdiger Scheible
Supplemental Figure 1. Highly Expressed Genes in the Hormone Series show low Coefficients of Variation due to high Scanning Sensitivity Resulting in Signal Saturation.
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Supplemental Text
The developmental series was analysed further to identify whether specific types of samples are responsible for the higher CV. When the data from seed samples (24 ATH1 arrays; samples #76-84 in Figure 1 for example) were excluded from the developmental set, the CV values for the Top100 genes decreased to 0.128 – 0.182 (cp. supplemental Table I), suggesting that the seed expression values increase the CV in general. Indeed, for 52 of the 58 genes present in both sets of 100 the CV decreased when the seed expression values were excluded, and for the other six genes CV was unchanged or increased only slightly (see supplemental Figure 2A). Extension of this analysis to the > 12,300 genes (called ‘present’ at a frequency of >80%, see above) shows that for over two thirds of the genes CV decreases when seed samples are omitted (supplemental Figure 2B).Pollen samples have very significant influence on the CV. For 65 of the 78 genes that are shared among the sets for the developmental series with or without pollen, the CV was decreased by omitting the pollen expression values. For the remaining 13 genes CV remained almost unchanged (supplemental Figure 2C). Removal of the pollen data leads to the appearance of 22 ‘new’ genes in the Top100 list. The CV for these genes also decreases markedly when pollen samples are excluded (supplemental Figure 2E, blue bars). Graphical painting of the developmental, relative expression patterns for the sixgenes with the largest decrease in CV (supplemental Figure 2F) demonstrates that they show very different expression in pollen (sample #73) compared to all other samples. Two (i.e. At5g51880 and At1g76940) have absolute expression levels (cp. mean values in supplemental Table I) that clearly suggest conserved expression across all developmental samples. In contrast, the other four genes have low absolute expression values, raising the question, whether they are indeed expressed at constant but low levels (as indicated by the high percentage of ‘present’ calls) across the developmental series, or whether they are pollen-specific genes. Less expected was the decrease of the CV for 18 of the 22 genes that were present in the Top100 list before pollen-subtraction (supplemental Figure 2E, red bars). The analysis of > 12,200 genes (called ‘present’ at a frequency of >80%) shows that removal of the pollen samples in fact leads to lower CVs for a very large majority of genes (supplemental Figure 2D).Exclusion of other sample subsets such as roots, i.e. #3, 9, 93-95, 98, 99, (supplemental Figure 3A, B) or flowers, i.e. #31-45, 53-59 (supplemental Figure 3C, D) from the developmental series did not lead to a marked increase in the percentages of genes that have a lower CV after sample subset omission. Indeed, omission of e.g. flower samples led to increased CV values for ~75% of the expressed genes (supplemental Figure 3D). This indicates that pollen and, to a lesser extent, seeds have transcriptomes that are very different from other tissues. A similar conclusion is reached by analysing the gene expression ratios in pollen and the other developmental samples (supplemental Figure 4A). Only a third (7431) of the ATH1 probe sets result in similar (<2-fold different) hybridization signals in pollen and the other developmental samples, whereas in flowers about 84% (19176) of the probe sets give similar signals (supplemental Figure 4B).
Supplemental Figure 2. Coefficients of Variation Predominantly Decrease after Omission of (A,B) Seed or (C,D) Pollen Samples from the Developmental Series.
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Supplemental Figure 2. continued
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Supplemental Figure 3. No Predominant Decrease of Coefficients of Variation after Omission of (A,B) Root or (C,D) Flower Samples from the Developmental Series.
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Supplemental Figure 4. Pollen (A) but not Flower Samples (B) display markedly changed Transcriptomes.
7431genes 19176 genes
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Supplemental Figure 5. Relative Expression Strengths of Additional (A) Beta-Tubulin, (B) Actin and (C) Poly-Ubiquitin Genes during Development.
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Coefficientof Variation
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Affymetrix Probe Set
Samples of Developmental Series
Panel Descriptions:
(A) Developmental series. A full description of samples 1-101 is available at http://www.weigelworld.org/resources/microarray/AtGenExpress/.
(B, C) Abiotic stress series for (B) shoots and (C) roots. A detailed description of the samples and plant growth conditions is available at http://web.uni-frankfurt.de/fb15/botanik/mcb/AFGN/atgenextable2.htm.
(D) Hormone series. Wild-type Col-0 seedlings were grown in liquid MS medium for seven days at 23°C treated with IAA, zeatin, GA3, ABA, MJ, ACC or BL for 30 minutes, 1 or 3 hours.
(E) Nutrient starvation and re-addition series. Wild-type Col-0 seedlings were grown as described in Scheible et al. (2004) and in Materials. Full nutrition (FN); sulfur, nitrogen, phosphorus or carbon starvation (-S, -N, -P, -C, respectively); 30 min or 3 hrs sulfate (SO4), nitrate (NO3), phosphate (PO4) or sucrose (suc) re-addition.
(F) Diurnal series / extended night / pgm mutant. Wild-type Col-0 or pgm mutant (light blue line) were grown and harvested as described in Bläsing et al. (2005).
(G) Light series. A detailed description of the samples is available at http://web.uni-frankfurt.de/fb15/botanik/mcb/AFGN/atgenextable3.htm.
(H) Biotic stress series. A detailed description of the samples is available at http://web.uni-frankfurt.de/fb15/botanik/mcb/AFGN/atgenextable3.htm.
Supplemental Figure 6. Comprehensive ATH1 GeneChip® Expression Profiles for 30 Traditional and Novel Housekeeping Genes.
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on V
alue
0
2000
4000
6000
8000
10000
12000
14000
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
2000
4000
6000
8000
10000
12000
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
2000
4000
6000
8000
10000
12000
14000
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
10000
20000
30000
40000
50000
FN -S30
min SO43h
SO4 -N30
min NO3
3h N
O3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
2000
4000
6000
8000
10000
12000
14000
16000
18000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
1000
2000
3000
4000
5000
6000
7000
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
2000
4000
6000
8000
10000
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At1g13440, GAPDH, Affymetrix Probe Set 259361_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
100
200
300
400
500
600
700
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
200
400
600
800
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
1000
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
1000
2000
3000
4000
5000
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
1000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
200
400
600
800
1000
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At5g25760, UBC, SIGMA-Aldrich house-keeping geneC3240, Affymetrix Probe Set 246862_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
100
200
300
400
500
600
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
1400
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
100
200
300
400
500
600
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
1000
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
500
1000
1500
2000
2500
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
100
200
300
400
500
600
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
20
40
60
80
100
120
140
160
180
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
50
100
150
200
250
300
350
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At5g44200, CBP20, SIGMA-Aldrich house-keeping geneC3115, Affymetrix Probe Set 249036_at
At1g13320, PDF2, Affymetrix Probe Set 259407_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
200
400
600
800
1000
1200
1400
1600
1800
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
200
400
600
800
1000
1200
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
1000
2000
3000
4000
5000
6000
7000
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
1000
1200
1400
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
100
200
300
400
500
600
700
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
200
400
600
800
1000
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
100
200
300
400
500
600
700
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
100
200
300
400
500
600
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
100
200
300
400
500
600
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
1000
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
500
1000
1500
2000
2500
3000
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
1000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
100
200
300
400
500
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
100
200
300
400
500
600
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At4g34270, TIP41-like gene, Affymetrix Probe Set 253287_at
At1g59830, PP2A gene, Affymetrix Probe Set 262909_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
200
400
600
800
1000
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
100
200
300
400
500
600
700
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
100
200
300
400
500
600
700
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
100
200
300
400
500
600
700
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
500
1000
1500
2000
2500
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
100
200
300
400
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
100
200
300
400
500
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
200
400
600
800
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
100
200
300
400
500
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
200
400
600
800
1000
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
100
200
300
400
500
600
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
500
1000
1500
2000
2500
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
1000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
50
100
150
200
250
300
350
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
200
400
600
800
1000
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At4g33380, expressed gene, Affymetrix Probe Set 253355_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
100
200
300
400
500
600
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
200
400
600
800
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
100
200
300
400
500
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
100
200
300
400
500
600
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
1000
2000
3000
4000
FN -S30
min SO43h
SO4 -N30
min NO3
3h N
O3 -P30
min PO43h
PO4 -C30
min Suc3h
Suc
E
0
100
200
300
400
500
600
700
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
100
200
300
400
500
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
100
200
300
400
500
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At2g28390, SAND gene family member,Affymetrix Probe Set 265256_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
200
400
600
800
1000
1200
1400
1600
1800
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
1400
1600
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
1400
1600
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
2000
4000
6000
8000
10000
FN -S30
min SO43h
SO4 -N30
min NO3
3h N
O3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
1000
1200
1400
1600
1800
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
200
400
600
800
1000
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
200
400
600
800
1000
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At4g26410‚ expressed gene, Affymetrix Probe Set 253959_at
At2g07190, hypothetical gene, Affymetrix Probe Set 266429_atExpression couldn‘t be confirmed by qPCR, nor were any cDNAs, ESTs, or MPSS signatures reported.
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
5
10
15
20
25
30
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
5
10
15
20
25
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
5
10
15
20
25
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
5
10
15
20
25
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
50
100
150
200
250
300
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
5
10
15
20
25
30
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
5
10
15
20
25
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
5
10
15
20
25
30
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
5
10
15
20
25
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
10
20
30
40
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
5
10
15
20
25
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
5
10
15
20
25
30
35
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
50
100
150
200
250
300
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
5
10
15
20
25
30
35
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
5
10
15
20
25
30
35
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
5
10
15
20
25
30
35
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At4g38070, bHLH transcription factor gene, Affymetrix Probe Set 253023_at ()
At2g32170‚ expressed gene, Affymetrix Probe Set 265697_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
100
200
300
400
500
600
700
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
100
200
300
400
500
600
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
200
400
600
800
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
1000
2000
3000
4000
5000
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
1000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
100
200
300
400
500
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
200
400
600
800
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
500
1000
1500
2000
2500
3000
3500
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
1000
2000
3000
4000
5000
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
500
1000
1500
2000
2500
3000
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
1000
2000
3000
4000
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
2000
4000
6000
8000
10000
12000
14000
16000
FN -S30
min SO43h
SO4 -N30
min NO3
3h N
O3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
1000
2000
3000
4000
5000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
500
1000
1500
2000
2500
3000
3500
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
500
1000
1500
2000
2500
3000
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At5g08290, YLS8, Affymetrix Probe Set 246006_at
At5g12240‚ expressed gene, Affymetrix Probe Set 250311_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
20
40
60
80
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
2
4
6
8
10
12
14
16
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
10
20
30
40
50
60
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
10
20
30
40
50
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
50
100
150
200
250
300
350
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
10
20
30
40
50
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
10
20
30
40
50
60
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
10
20
30
40
50
60
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At1g62930, PPR-repeat gene, Affymetrix Probe Set 261095_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
20
40
60
80
100
120
140
160
180
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
50
100
150
200
250
300
350
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
50
100
150
200
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
50
100
150
200
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
100
200
300
400
500
600
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
50
100
150
200
250
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
20
40
60
80
100
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
20
40
60
80
100
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At3g53090, UPL7, Affymetrix Probe Set 251998_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
50
100
150
200
250
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
50
100
150
200
250
300
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
50
100
150
200
250
300
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
100
200
300
400
500
600
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
1400
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
100
200
300
400
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
50
100
150
200
250
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
50
100
150
200
250
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
10
20
30
40
50
60
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
5
10
15
20
25
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
5
10
15
20
25
30
35
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
5
10
15
20
25
30
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
50
100
150
200
250
300
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
10
20
30
40
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
10
20
30
40
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
10
20
30
40
50
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At1g47770, hypothetical gene, Affymetrix Probe Set 261732_atExpression couldn‘t be confirmed by qPCR, nor were any cDNAs, ESTs, or MPSS signatures reported.
At3g01150, PTB, Affymetrix Probe Set 259280_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
100
200
300
400
500
600
700
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
100
200
300
400
500
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
100
200
300
400
500
600
700
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
500
1000
1500
2000
2500
3000
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
200
400
600
800
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
100
200
300
400
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
100
200
300
400
500
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
20
40
60
80
100
120
140
160
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
020406080
100120140160180200
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
50
100
150
200
250
300
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
50
100
150
200
250
300
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
1400
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
50
100
150
200
250
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
20
40
60
80
100
120
140
160
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
20
40
60
80
100
120
140
160
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At4g16320‚ expressed gene, Affymetrix Probe Set 245491_at
At3g32260, hypothetical gene, Affymetrix Probe Set 256640_atExpression couldn‘t be confirmed by qPCR, nor were any cDNAs, ESTs, or MPSS signatures reported.
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
10
20
30
40
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
5
10
15
20
25
30
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
10
20
30
40
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
20
40
60
80
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
50
100
150
200
250
300
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
10
20
30
40
50
60
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
10
20
30
40
50
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
10
20
30
40
50
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At5g15710, F-box gene, Affymetrix Probe Set 246515_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
50
100
150
200
250
300
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
020406080
100120140160180200
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
50
100
150
200
250
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
50
100
150
200
250
300
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
200
400
600
800
1000
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
50
100
150
200
250
300
350
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
020406080
100120140160180200
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
20
40
60
80
100
120
140
160
180
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At4g27960, At5g53300, UBC9 & UBC10,Affymetrix Probe Set 253826_s_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
2000
4000
6000
8000
10000
12000
14000
16000
18000
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
2000
4000
6000
8000
10000
12000
14000
16000
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
2000
4000
6000
8000
10000
12000
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
2000
4000
6000
8000
10000
12000
14000
16000
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
10000
20000
30000
40000
50000
FN -S30
min SO43h
SO4 -N30
min NO3
3h N
O3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
2000
4000
6000
8000
10000
12000
14000
16000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
2000
4000
6000
8000
10000
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
2000
4000
6000
8000
10000
12000
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At5g46630, clathrin adaptor complex medium subunit geneAffymetrix Probe Set 248858_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
200
400
600
800
1000
1200
1400
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
1400
1600
1800
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
200
400
600
800
1000
1200
1400
1600
1800
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
1000
2000
3000
4000
5000
6000
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0200400600800
100012001400160018002000
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
200
400
600
800
1000
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
200
400
600
800
1000
1200
1400
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GST
water
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At5g55840, PPR repeat gene, Affymetrix Probe Set 248024_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
10
20
30
40
50
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
AEx
pres
sion
Val
ue
0
10
20
30
40
50
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
10
20
30
40
50
60
70
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
10
20
30
40
50
60
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
50
100
150
200
250
300
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
20
40
60
80
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
G
Expr
essi
on V
alue
0
5
10
15
20
25
30
35
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
10
20
30
40
50
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
At1g58050, helicase-like gene,Affymetrix Probe Set 245863_s_at
0h 4h 8h 12h
16h
20h 0h 2h 4h 6h 8h 24h
48h
F
0
20
40
60
80
100
120
140
160
180
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 31 32 33 34 35 36 37 39 40 41 42 43 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 73 76 77 78 79 81 82 83 84 87 89 90 91 92 93 94 95 96 97 98 99 100
101
A
Expr
essi
on V
alue
0
50
100
150
200
250
300
350
0min
15m
in30
min 1h 3h 4h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
30m
in 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h 6h 12h
24h
15m
in30
min 1h 3h
3h+1
h3h
+3h
3h+9
h3h
+21h
B
Expr
essi
on V
alue
0
20
40
60
80
100
120
140
controls cold (4°C) osmotic salt drought genotoxic oxidative UV-B wounding heat (38°C)+ recovery
C
Expr
essi
on V
alue
0
100
200
300
400
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
30m
in 1h 3h30
min 1h 3h
D
Expr
essi
on V
alue
0
100
200
300
400
500
FN -S
30min SO4
3h SO4 -N
30min N
O33h
NO3 -P
30min PO4
3h PO4 -C
30min Suc
3h Suc
E
0
50
100
150
200
250
300
350
mock ABA ACC BL GA3 IAA MJ zeatin
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
45m
in 4h45
min 4h
GEx
pres
sion
Val
ue
0
5
10
15
20
25
30
35
0h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 2h 6h 24h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 1h 4h 6h 12h
24h 6h 12h
24h
H
0
10
20
30
40
50
60
70
Dark
10 µE Far Red
1' Red +Dark
10µE Red
10µE Blue
5' UV-A +Dark
5' UV-A/B + Dark
10µE White DC3000
avrRpm1
DC3000 hrcC-
phaseolicola
10mM MgCl 2
1µM GSTwater
Ca / Mg
10 µM HrpZ
1 µM GST-NPP1
1 µM Flg22
Phytophthora
infestans
100 µg/ml L
PS + -
Extended Night
Supplemental Figure 7. geNORM Ranking of 27 Housekeeping Genes based on ATH1 Data from Experimental Series A-H (see Legend to supplemental Figures 6.1-6.30)
<-- least stable most stable -->
At5g12
240
At1g58
050
At5g12
250
At3g18
780
At5g44
200
At5g09
810
At4g33
380
At3g53
090
At1g59
830
At5g60
390
At1g13
440
At5g25
760
At5g55
840
At4g16
320
At4g38
070
At5g46
630
At3g01
150
At5g15
710
At1g62
930
At2g28
390
At5g08
290
At4g34
270
At4g26
410
At4g27
960
At2g32
170
At4g05
320
At1g13
320Av
erag
e Ex
pres
sion
Sta
bilit
y M
0,0
0,2
0,4
0,6
0,8
1,0
1,2
1,4
AGI Annotation SelectionCriterium 5’ 3’ Primer Sequences (F/R) Primer
Efficiencyb
Amplicon Length
(bp) Amplicon
TM (°C) Distance from 3’ end C
AT1G13440 GAPDH T a1 a4 TTGGTGACAACAG^GTCAAGCA AAACTTGTCGCTCAATGCAATC 0.94±0.07 62 77.4 326
AT3G18780 ACT2 T G CTTGCACCAAGCAGCATGAA CCGATCCAGACACTGTACTTCCTT 0.91±0.01 68 79.8 561
AT4G05320a UBQ10 T a1 A2 a4 c h
GGCCTTGTATAATCCCTGATGAATAAG AAAGAGATAACAGGAACGGAAACATAGT 0.91±0.02 61 70.4 244
AT5G25760 UBC T a2 G CTGCGACTCAG^GGAATCTTCTAA TTGTGCCATTGAATTGAACCC 0.89±0.01 61 73.2 173
AT5G60390 EF-1α T a1 a4 C G h
TGAGCACGCTCTTCTTGCTTTCA GGTGGTGGCATCCAT^CTTGTTACA 0.95±0.01 76 78.7 1221
AT1G13320 PP2A subunit PDF2
A1 A2 A3 A4 B C d e G h
TAACGTGGCCAAAATGATGC GTTCTCCACAAC^CGCTTGGT 0.93±0.01 61 76.0 288
AT1G47770 Hypothetical protein B C E GTTCAATAAAATGGCGCATCTTG
GAAAAGGTGCAAAACGATCTCAC NA NA NA 129
AT1G58050 Helicase A1 A2 A3 A4
CCATTCTACTTTTTGGCGGCT TCAATGGTAACTGATCCACT^CTGATG 0.85±0.01 61 73.0 184d
AT1G62930 PPR gene c d E G H GAGTTGCGGGTTTGTTGGAG CAAGACAGCATTTCCAGATAG^CAT 0.88±0.01 61 75.0 129d
AT2G07190 Hypothetical protein
A1 A2 A3 A4 B C E
CCGTCCAATCCAACAGATCAG CGTCATCTAAAGACATTAGGTCCTGTAC NA NA NA 209
AT2G28390 SAND family A1 A2 A3 A4 c e G h
AACTCTATGCAGCATTTGATCCACT TGATTGCATATCTTTATCGCCATC 0.83±0.01 61 75.8 323
AT2G32170 expressed A1 A2 A3 A4 B c d E
ATCGAGCTAAGTTTGGAGGATGTAA T^CTCGATCACAAACCCAAAATG 0.80±0.01 61 74.1 293
AT3G01150Polypyrimidine-
tract-binding protein
A1 A2 A3 A4 d e g H
GATCTGAATGTTAAG^GCTTTTAGCG GGCTTAGATCAGGAAGTGTATAGTCTCTG 0.74±0.01 61 72.0 332
AT3G32260 Hypothetical protein
A1 A2 A3 A4 B c E
CTGTTTGCCGAAGTTCCAGAGT TTAAATCAGCAAGAACGTCGGATA NA NA NA 293
AT3G53090 Ubiquitin-transferase a1 B E TTCAAATACTTGCAGCCAACCTT
CCCAAAGAGAGGTATCACAAGAGAC^T 0.73±0.01 61 72.3 431
AT4G26410 expressed A1 A2 A3 a4 D E g H
GAGCTGAA^GTGGCTTCCATGAC GGTCCGACATACCCATGATCC 0.91±0.01 81 77.4 242
AT4G27960 UBC9 A1 a4 b c E G h
TCACAATTTCCAAG^GTGCTGC TCATCTGGGTTTGGATCCGT 0.88±0.01 61 74.1 330
AT4G33380 expressed A1 A2 A3 A4 D
TTGAAAATTGGAGTACCGTACCAA TCCCTCGTATACATCTGGCCA 0.81±0.01 61 73.2 122
AT4G34270 TIP41-like A1 A2 A3 A4 b C e
GTGAAAACTGTTGGAGAGAAGCAA TCAACTGGATACCCTTT^CGCA 0.86±0.01 61 73.4 388
AT4G38070 bHLH a1 a4 B C E
GAAAGCAAAGGCGGTGAGAG CAAGGCACACTTGGTTCTTCC 0.75±0.02 61 77.3 135d
AT5G08290 mitosis protein YLS8
A1 A3 a4 B F G h
TTACTGTTTCGGTTGTTCTCCATTT CACTGAATCATGTTCGAAGCAAGT 0.81±0.01 66 71.3 31
AT5G12240 expressed A1 A3 a4 B C
AGCGGCTGCTGAGAAGAA^GT TCTCGAAAGCCTTGCAAAATCT 0.85±0.01 63 76.9 286
AT5G15710 F-box protein a1 b C g TTTCGGCTGAGAGGTTCGAGT GATTCCAAGACGTAAAGCAGATCAA 0.91±0.01 63 74.0 359
AT5G46630 clathrin adaptor complex subunit
A1 A2 A3 A4 C
TCGATTGCTTGGTTTGGAAGAT GCACTTAGCGTGGACTCTGTTTG^ATC 0.86±0.01 61 75.0 438
AT5G55840 PPR gene A1 A2 A3 A4 b c E
AAGACAGTGAAGGTGCAACCTTACT AGTTTTTGAGTTGTATTTGTCAGAGAAAG 0.74±0.01 61 74.1 348d,e
Supplemental Table II. Traditional and Novel Housekeeping Genes selected for qPCR Validation, and Specifications of Primers and Amplicons.
a primers do not work on C24 cDNA; primers recognize all known splice variants, b Mean ± SE, n=60; cDistance from end of longest known 3’ UTR ; d Distance from electronically annotated stop codon; e No experimentally verified gene model available; NA = no amplification; ^ position of an exon-exon junction. Explanation of selection criteria abbreviations: traditional housekeeping gene (T) ; Top100 member of (A1) complete developmental series, (A2) developmental series without seed samples, (A3) developmental series without pollen sample, (A4) developmental series without seed and pollen samples, (B) shoot abiotic stress series, (C) root abiotic stress series, (D) Hormone series, (E) nutrient stress series, (F) diurnal series, (G) light series, (H) biotic stress series. Letters were chosen according to the panel descriptions in supplemental Figures 6.1 – 6.30. Small letters indicate the presence in the extended Top500 list of each series.
Supplemental Table III. cDNA Samples used for Real-Time PCR Validation of Housekeeping Genes.
cDNA Pool Plant Material used for RNA / cDNA Preparation Wild-type
EcotypeGAPDH 5'/3' Ratio
Mean ± SD a
1 9 days old liquid culture grown (LC) seedlings, full nutrients (FN) Col-0 0,62 ± 0,07 2 9 days old LC seedlings, FN, 3 hours 100mM NaCl Col-0 0,58 ± 0,16 3 9 days old LC seedlings, FN, 3 hours 200mM Mannitol Col-0 0,66 ± 0,10 4 9 days old liquid culture grown sulphur-depleted LC seedlings Col-0 0,50 ± 0,05 5 9 days old carbohydrate depleted LC seedlings Col-0 0,13 ± 0,02 6 9 days old phosphorus-depleted LC seedlings Col-0 0,70 ± 0,03 7 Shoots of 14 days old plants grown on vertical agar plates Col-0 0,83 ± 0,05 8 Roots of 14 days old plants grown on vertical agar plates Col-0 0,61 ± 0,19 9 Shoot apices of 4 weeks old soil-grown plants Col-0 0,59 ± 0,03 10 Rosette leaves of 4 weeks old soil-grown plants Col-0 0,58 ± 0,06 11 Rosette leaves of 6 weeks old soil-grown plants Col-0 0,25 ± 0,03 12 Cauline leaves of 5 weeks old soil-grown plants Col-0 0,39 ± 0,03 13 Stems of 5 weeks old soil-grown plants Col-0 0,56 ± 0,04 14 Flowers of 6 weeks old soil-grown plants Col-0 0,39 ± 0,01 15 Siliques of 6 weeks old soil-grown plants Col-0 0,61 ± 0,09 16 10 days old LC seedlings, no exogenous hormone addition C24 0,24 ± 0,07 17 10 days old LC seedlings, 3 hours 2,4-dichlorophenoxyacetic acid C24 0,64 ± 0,01 18 6 weeks old plants, cold-unacclimated Col-0 0,36 ± 0,01 19 6 weeks old plants, cold-acclimated Col-0 0,93 ± 0,02 20 Seeds of 8 week old soil-grown plants Col-0 0,41 ± 0,06
a mean values ± standard deviation (SD) of three measurements.
Supplemental Figure 8. Dissociation Curves for PCR Products amplified with the Primer Pairs shown in supplemental Table II.
At3g18780
At1g13320 At1g58050 At2g32170At2g28390
At4g26410 At4g27960
At4g33380 At4g34270 At4g38070 At5g15710
At5g25760 At5g46630
At5g55840 At5g60390
At4g05320
At1g13440
At1g62930
At3g01150 At3g53090
At5g08290 At5g12240
LjLb2 At1g13440 (5‘)
GTGAAAACTGTTGGAGAGAAGCAACATTTCAGTCTCTATCTGCGAAAGGGTATCCAGTTGA
At4g34270
At5g15710
At4g26410
At2g28390
FWD Primer REV PrimerExpected Amplicon
TTTCGGCTGAGAGGTTCGAGTGTTTTGGACAAGATAATTTGATCTGCTTTACGTCTTGGAATC
GAGCTGAAGTGGCTTCCATGACTTCGCTTTTGAGACAGAAGAGGATTGCATTGAACAAAAGGATCATGGGTATGTCGGACC
AACTCTATGCAGCATTTGATCCACTTGCAGACAAGGCGATGGCGATAAAGATATGCAATCA
Supplemental Figure 10.
Sequence Electropherogramsof four Amplicons.
Supplemental Figure 9.
Agarose Gel Analysis of additional PCR amplicons.
At5
g082
90
At5
g558
40
At2
g321
70
At4
g279
60
At4
g342
70
At1
g580
50
At4
g264
10
50
100
150
At1
g477
70
At3
g011
50
At1
g629
30
At5
g157
10
At5
g122
40
150
10050
Supplemental Figure 11. geNorm-Ranking of 22 Housekeeping Genes based on qPCR Data.
Data from 18 cDNA pools (samples from C24 ecotype background were omitted) are shown for traditional (black bars) and novel housekeeping genes (white bars). A low value for the average expression stability M, as calculated by geNorm software, indicates a more stable expression..
<-- least stable most stable -->
AT5G60
390
AT3G18
780
AT4G38
070
AT1G62
930
AT4G05
320
AT4G27
960
AT1G58
050
AT1G13
440
AT1G13
320
AT5G12
240
AT3G01
150
AT3G53
090
AT2G32
170
AT5G55
840
AT5G08
290
AT5G15
710
AT4G26
410
AT5G25
760
AT4G33
380
AT5G46
630
AT4G34
270
AT2G28
390
Aver
age
Expr
essi
on S
tabi
lity
M
0,0
0,5
1,0
1,5
