The Arabidopsis Functional Genomics Network

AtGenExpress

Overview
Developmental samples
Stress treatments
GARNET: Light treatments, Pathogen infection, Base line experiments

 

Table 3: GARNET

Series 1 (samples 1-16, taken as triplicates): Light treatments

Series 2 (samples 17-46, taken as triplicates): Pathogen infection, base line experiments

Series 1 (samples 1-16): Growth conditions: Sterilized seeds will be stratified at 4°C for 3 days, exposed to white light for 2 h to induce germination, and grown on MS agar plates (0.9 % agar) without sugar in total darkness for 4 days at 22°C. Seedlings will then be transferred to the light conditions described beelow for 1 h (to identify early induced genes) and 4 hrs (maximum expression of the first initial light response of most target genes), respectively. All light treatments will be done in parallel to minimize the number of dark controls. All samples will be done in triplicate and only with shoots.

Sample

Organs

Treatments (after 4 days of continuous darkness)

Group

1.       

Shoots

30 min darkness (handling control)

Kretsch

2.       

Shoots

4 hr darkness (handling control)

Kretsch

3.       

Shoots

30 min far-red light (716 nm DAL interference filter, 10 µmol m-2 sec-1)

Kretsch

4.       

Shoots

4 hrs far-red light (716 nm DAL interference filter, 10 µmol m-2 sec-1)

Kretsch

5.       

Shoots

30 min red light (KG 65 filter, lambda (max)= 650 nm, 10 µmol m-2 sec-1)

Kretsch

6.       

Shoots

Continuous darkness interrupted by a 1 min saturating red light pulse (KG 65 filter, lambda (max) = 650 nm, 50 µmol m-2 sec-1) followed by 30 min darkness

Kretsch

7.       

Shoots

Continuous darkness interrupted by a 1 min saturating red light pulse (KG 65 filter, lambda (max) = 650 nm, 50 µmol m-2 sec-1) followed by 4 hrs darkness

Kretsch

8.       

Shoots

4 hrs red light (KG 65 filter, lambda (max) = 650 nm, 10 µmol m-2 sec-1

Kretsch

9.       

Shoots

30 min blue light (453 nm DAL interference filter, 10 µmol m-2 sec-1);

Kretsch

10.   

Shoots

4 hrs blue light (453 nm DAL interference filter, 10 µmol m-2 sec-1);

Kretsch

11.   

Shoots

Continuous darkness interrupted  by a 5 min UV-A/327 nm cut-off light pulse followed by 30 min darkness

Kretsch

12.   

Shoots

Continuous darkness interrupted  by a 5 min UV-A/327 nm cut-off light pulse followed by 4 hrs darkness

Kretsch

13.   

Shoots

Continuous darkness interrupted  by a 5 min UV-B/305 nm cut-off light pulse followed by 30 min darkness

Kretsch

14.   

Shoots

Continuous darkness interrupted  by a 5 min UV-B/305 nm cut-off light pulse followed by 4 hrs darkness

Kretsch

15.   

Shoots

30 min white light (Osram Xenophot longlife lamps without filters, 10 µmol m-2 sec-1)

Kretsch

16.   

Shoots

4 hr white light (Osram Xenophot longlife lamps without filters, 10 µmol m-2 sec-1)

Kretsch


 

Series 2 (samples 17-46) Base line experiment for pathogen infection. Growth and infection conditions: Seeds will be stratified for 3 days at 4°C and sown on soil. Plants are grown at 22°C under a 8/16 hour light/dark regime in Percival growth chambers. All pathogen treatments will be performed on leaves of 5-weeks old plants. Bacterial infiltrations will be performed with 10-8 cfu/ml in 10 mM MgCl2. 5x105 spores of Phytophthora infestans in water will be applied to leaf surfaces. LPS (100 µg/ml), HrpZ (1 µM), NPP1 (2 µM) or flagellin (1 µM flg22) will be applied in water. Experiments should be performed in triplicate at the time points indicated below. 

17-20

Leaves

Water control taken at 0, 2, 6 and 24 hrs of treatment

Nürnberger

21-23

Leaves

Pseudomonas syringae pv tomato (Pst) DC3000 2/6/24 hrs p.i.

Nürnberger

24-26

Leaves

Pst AvrRpm1, 2/6/24 hrs p.i.

Nürnberger

27-29

Leaves

Pst DC3000 hrcC-, 2/6/24 hrs p.i.

Nürnberger

30-32

Leaves

Pseudomonas syringae pv phaseolicola (nonhost) 2/6/24 hrs p.i.

Nürnberger

33-35

Leaves

MgCl control 2/6/24 hrs p.i.

Nürnberger

36-38

Leaves

Phytophthora infestans 2/6/24 hrs p.i.

Nürnberger

39-40

Leaves

LPS, 1 and 4 hrs p.i.

Nürnberger

41-42

Leaves

HrpZ, 1 and 4 hrs p.i.

Nürnberger

43-44

Leaves

NPP1, 1 and 4 hrs p.i.

Nürnberger

45-46

Leaves

Flg22, 1 and 4 hrs p.i.

Nürnberger

 

 

Coordinator: Lutz Nover, Detlef Weigel and Thomas Altmann