ath1 genome array  (Thermo Fisher)

Journal: Plant Physiology

Article Title: The transcription factor ORA59 represses hypoxia responses during Botrytis cinerea infection and reoxygenation

doi: 10.1093/plphys/kiae677

Figure Lengend Snippet: Expression of hypoxia-responsive genes during Botrytis infection A) bar plot of log 2 fold change (log 2 FC) expression values for core anaerobic genes after 48 h from the inoculation with Botrytis. Genes are sorted in descending order, with a dashed line indicating the threshold of log 2 FC = 1, representing significantly upregulated genes in adult Arabidopsis plants rosette 48 h post infection. B) Expression analysis of Group B core anaerobic genes after 48 h from infection with Botrytis and upon low-oxygen stresses. The treatments were 4 h of gaseous hypoxia and 4 h of submergence. Data ( n = 5) are relative to the mock group for Botrytis and to the air group for hypoxia/submergence treatments, both set to 1. C) Expression analysis of Group A core anaerobic genes after 48 h from infection with Botrytis and upon low-oxygen stress. The treatments were as in panel B) . Statistically significant differences in B) and C) are indicated by the asterisks (Student t -test, unpaired comparison was used for mock to Botrytis comparison, one-way ANOVA followed by Dunnett's multiple comparisons test was used for air to hypoxia and submergence comparisons. * = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). Lines within the boxes indicate the median, while the bottom and top of each box denote the first and third quartile, respectively, the dots represent the single data points and whiskers denote the min/max values.

Article Snippet: Affymetrix GeneChip Arabidopsis ATH1 Genome Arrays were used to process RNA, as previously described ( ).

Techniques: Expressing, Infection, Comparison

Journal: Plant Physiology

Article Title: The transcription factor ORA59 represses hypoxia responses during Botrytis cinerea infection and reoxygenation

doi: 10.1093/plphys/kiae677

Figure Lengend Snippet: Effects of jasmonate treatments on the expression of HRGs . A) Time-course expression analysis of jasmonate-responsive marker genes. Statistically significant differences are indicated by asterisks after Kruskal–Wallis test followed by Dunn's multiple comparisons test (* = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). Data ( n = 6) are relative to time 0 group set to 1. B) Expression analysis of Group B and A core anaerobic genes in adult Arabidopsis plants rosette pretreated with 50 µM MeJA for 7 h and then submerged in the dark for 2 h. Different letters indicate significant difference ( P < 0.05) after two-ways ANOVA followed by Tukey's post-hoc test. Data ( n = 6) are relative to the Col-0 control (air) not pretreated with 50 μM MeJA, set to 1. C) Representative images of LUC activity in pADH:LUC and 5xHRPE:LUC plants after 7 h incubation with 50 µM MeJA and 2 h of gaseous hypoxia. The scale bar is 1 cm. D) Quantification of LUC activity (counts per second, CPS) of pADH:LUC and 5xHRPE:LUC reporter lines showed in C) ( n ≥ 5). Quantification of CPS was carried out using the IndiGO software, by selecting a fixed area around the plants. Statistically significant differences are indicated by the asterisks (Student t -test, unpaired comparison; * = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). Lines within the boxes indicate the median, while the bottom and top of each box denote the first and third quartile, respectively, the dots represent the single data points and whiskers denote the min/max values.

Article Snippet: Affymetrix GeneChip Arabidopsis ATH1 Genome Arrays were used to process RNA, as previously described ( ).

Techniques: Expressing, Marker, Control, Activity Assay, Incubation, Software, Comparison

Journal: Plant Physiology

Article Title: The transcription factor ORA59 represses hypoxia responses during Botrytis cinerea infection and reoxygenation

doi: 10.1093/plphys/kiae677

Figure Lengend Snippet: ORA59 interacts with the ERFVIIs. A) Western blot analysis of RAP2.3 3xHA abundance in adult Arabidopsis plants leaves 48 h post infection with Botrytis followed by the quantification of the signal carried out using ImageLab TM software (BioRad). 40 μg of protein was loaded for each sample. Four biological replicates are shown. B) Expression analysis of ORA59 in Arabidopsis rosettes 48 h post infection. Data ( n = 6) are relative to the mock group set to 1. Statistically significant differences are indicated by the asterisks (Student t -test, unpaired comparison; * = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). C) Yeast two-hybrid assay between the 5 ERFVIIs and ORA59 (N-terminal 60 amino acids truncated, ORA59Δ1–60). D) As in C) , followed by a β-galactosidase assay (CPRG) test ( n = 4). E) Split-LUC assay of ORA59 fused to the N-terminal part of the firefly LUC and the ERFVIIs members (and negative control, HOL2) fused to the C-terminal part of the firefly LUC each ( n = 6). For D) and E) , statistically significant differences are indicated by asterisks after a one-way ANOVA followed by Dunnett's multiple comparisons test (* = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). Lines within the boxes indicate the median, while the bottom and top of each box denote the first and third quartile, respectively, the dots represent the single data points and whiskers denote the min/max values.

Article Snippet: Affymetrix GeneChip Arabidopsis ATH1 Genome Arrays were used to process RNA, as previously described ( ).

Techniques: Western Blot, Infection, Software, Expressing, Comparison, Y2H Assay, Negative Control

Journal: Plant Physiology

Article Title: The transcription factor ORA59 represses hypoxia responses during Botrytis cinerea infection and reoxygenation

doi: 10.1093/plphys/kiae677

Figure Lengend Snippet: ORA59 is a repressor of HRGs expression. A) Expression analysis of Group B and A core anaerobic genes in Col-0 and ora59 RNAi adult Arabidopsis rosette 48 h after inoculation with Botrytis. Data ( n = 6) are relative to the Col-0 mock set to 1. B) Expression of Group B and A core anaerobic genes in Col-0 protoplasts transformed with 35S:ORA59 and treated with gaseous hypoxia for 2 h. Data ( n = 4) are relative to the Col-0 untransformed control (air) set to 1. For A) and B) , different letters indicate significant difference ( P < 0.05) after two-ways ANOVA followed by Tukey's post-hoc test. C) Expression of ADH in erfVII protoplasts treated with gaseous hypoxia for 2 h. Protoplasts were transformed with each of the ERFVIIs singularly, under the control of the cauliflower mosaic virus 35S promoter, and with or without ORA59 driven by the same promoter. Statistically significant differences are indicated by asterisks after multiple unpaired t -test (* = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). Data ( n = 4) are relative to the erfVII untransformed group, set to 1. D) Inhibition of ERFVII-dependent expression of ADH by ORA59 in erfVII protoplasts. Data are the same as in and are mean ( n = 4) ±SD. Statistically significant differences are indicated by asterisks after a one-way ANOVA followed by Dunnett's multiple comparisons test (* = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). In box-plots, lines within the boxes indicate the median, while the bottom and top of each box denote the first and third quartile, respectively, the dots represent the single data points and whiskers denote the min/max values.

Article Snippet: Affymetrix GeneChip Arabidopsis ATH1 Genome Arrays were used to process RNA, as previously described ( ).

Techniques: Expressing, Transformation Assay, Control, Virus, Inhibition

Journal: Plant Physiology

Article Title: The transcription factor ORA59 represses hypoxia responses during Botrytis cinerea infection and reoxygenation

doi: 10.1093/plphys/kiae677

Figure Lengend Snippet: Production of ethanol is detrimental or Arabidopsis tolerance to Botrytis. A) Tolerance after Botrytis infection in Col-0, adh , pdc1pdc2 double mutants. The images show the lesion area 48 h after the inoculation. The scale bar is 1 cm. B) Percentage of lesion area normalized on the leaf size presented in a boxplot. Each dot represents the average value of a plant ( n = 12) (5 different infected leaves on each plant). C) Expression analysis in adult Arabidopsis rosette of B. cinerea actin A ( BcActinA ) as marker for the presence of the fungi. Data ( n = 6) are relative to Col-0 infected group set to 100 for BcActinA . For B) and C) , statistically significant differences are indicated by asterisks after a one-way ANOVA followed by Dunnett's multiple comparisons test (* = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). D) Transgenic line expressing GUS under the control of the ADH promoter ( pADH:GUS ). GUS staining was performed 48 h after the inoculation with Botrytis. The scale bar is 0.5 cm. E) Tolerance after Botrytis infection in Col-0 plants. Spores used in the infection were resuspended in either PDB (Control) or PDB added with 100 m M Ethanol. The images show the lesion area 48 h after the inoculation. The scale bar is 1 cm. F) Percentage of lesion area normalized on the leaf size presented in a boxplot. Each dot represents the average value of a plant ( n ≥ 9) (5 different infected leaves on each plant). G) Expression analysis in adult Arabidopsis rosette of B. cinerea actin A ( BcActinA ) as marker of the fungi presence. Data ( n = 9) are relative to Col-0 infected group set to 100 for BcActinA . For F) and G) , statistically significant differences are indicated by the asterisks (Student t -test, unpaired comparison; * = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). Lines within the boxes indicate the median, while the bottom and top of each box denote the first and third quartile, respectively, the dots represent the single data points and whiskers denote the min/max values.

Article Snippet: Affymetrix GeneChip Arabidopsis ATH1 Genome Arrays were used to process RNA, as previously described ( ).

Techniques: Infection, Expressing, Marker, Transgenic Assay, Control, Staining, Comparison

Journal: Plant Physiology

Article Title: The transcription factor ORA59 represses hypoxia responses during Botrytis cinerea infection and reoxygenation

doi: 10.1093/plphys/kiae677

Figure Lengend Snippet: The interplay between the ERFVIIs and ORA59 influences the reoxygenation phase after hypoxia. A) Expression analysis of ORA59 in Arabidopsis 5-d old seedlings during recovery in air for 5′ (R5′), 15′ (R15′), 30′ (R30′), and 1 h (R1) after 4 h of gaseous hypoxia (H4). Data ( n = 5) are relative to aerobic (T0) seedlings set to 1. B) LUC activity in 35S:RAP2.12 1–28 -LUC 7-d old seedling during post-hypoxia recovery in air for 1 h (R1), 2 h (R2), and 3 h (R3). For A) and B) , statistically significant differences ( n = 96) are indicated by the asterisks after a one-way ANOVA followed by Dunnett's multiple comparisons test (* = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). C) Expression analysis of Group B and A core anaerobic genes during recovery after 4 h of gaseous hypoxia in Col-0 and ora59 RNAi 5-d old seedlings. Curved lines represent polynomial fits passing through the mean ( n = 6) of each group of observations, providing a visual representation of the trend within each genotype. Statistically significant differences are assessed using two-ways ANOVA followed by Sidak's multiple comparisons test (* = P < 0.05; ** = P < 0.01; *** = P < 0.001; **** = P < 0.0001). Lines within the boxes indicate the median, while the bottom and top of each box denote the first and third quartile, respectively, the dots represent the single data points and whiskers denote the min/max values.

Article Snippet: Affymetrix GeneChip Arabidopsis ATH1 Genome Arrays were used to process RNA, as previously described ( ).

Techniques: Expressing, Activity Assay