1
Mariam Awlia 1 , Arianna Nigro 2 , Jiří Fajkus 3 , Martin Trtílek 3 , Diana Santelia 2 , Mark A. Tester 1 , Magdalena M. Julkowska 1 and Klára Panzarová 3 1 King Abdullah University of Science and Technology (KAUST), Thuwal, Saudi Arabia; 2 Institute of Plant Biology, University of Zurich, Switzerland; 3 PSI (Photon Systems Instruments), Czech Republic Soil salinity is one of the main stress factors severely affecting the agriculture land in global scale and causing significant reduction of plant growth and yield. To enhance our understanding of the early responses to salinity, we designed an experimental protocol based on automated integrative analysis of photosynthetic performance, growth analysis and color index analysis at the onset and early phase of salinity stress response in Arabidopsis thaliana ecotypes grown in soil. Here we show that the developed experimental procedure allows to analyse dynamically structural and physiological phenotypes very early upon stress imposition. Results for two accessions Col-0 and C24 are shown. C24 was previously described for increased salt tolerance. Salinity significantly and rapidly affected photosynthetic performance and impacted growth dynamics of Arabidopsis plants at different stages of stress response. PLANT PHENOTYPING RESEARCH CENTER Screening tools for identification of traits contributing to salinity tolerance in Arabidopsis PLANTSCREEN TM AUTOMATED PHENOTYPING SYSTEMS www.plantphenotyping.com www.psi.cz The mission of the PSI Plant Phenotyping Research Center (PPRC) is to provide state-of-art infrastructure for plant cultivation and automated high-throughput phenotyping of wide range of plant traits. We offer access to cutting edge instruments and provide professional support of highly skilled technical and scientific personnel. PPRC infrastructure is available for use by visiting scientists and on fee-for- service basis for a wide range of phenotyping experiments. PPRC operates high-end walk-in growth chambers for precise growth of plants and PlantScreen TM platforms for automated phenotyping of small and mid- size plants in controlled environment (e.g. turfgrass, Arabidopsis thaliana) and for cultivation and monitoring of larger crop plants up to 1.5 meter in height. Chlorophyll fluorescence kinetic imaging and analysis of photosynthetic performance Max light adapted fluorescence (Fp) PAM light LED panel for kinetic imaging F O , F M , F V , F O' , F M' , F V' , F t Max quantum efficinecy Fv/Fm Photochemical quenching Non-photochemical quenching Vitality index F V' /F M' , Phi PSII , qN, qP Fluorescence kinetics measurement Mask application and bckgr substraction Fluorescence quenching kinetics Original image Barrel distortion correction Color segmentation Background subtraction Mask detection Area Perimeter Roundness Compactness Leaf development tracking Greening index RLGR PPRC is situated in countryside next to Brno in Czech Republic, city where Johann Gregor Mendel lived and worked. For more information contact us at [email protected]. Automated phenotyping of up to 320 small- and mid-size scale plants in controlled environment in PlantScreen TM Compact System. Automated phenotyping of 270 plants up to 1.5 in height in greenhouse environment in PlantScreen TM Modular System. Large-scale walk in chambers for highly precise plant cultivation. Rationale Phenotyping Protocol Arabidopsis thaliana Col-0 and C24 accessions were grown in 12h-12h light conditions under cool-white LED illumination of 150 μmol/m 2 /s in FS-WI Chamber (PSI). Before the salt imposition plants were automatically weighed and watered using PlantScreen TM Phenotyping System to adjust soil moisture to 60% of soil water capacity (A). At 21 days after stratification (DAS) plants were treated with 250 mM NaCl solution for one hour, ensuring saturation of the soil with the solution. The effective NaCl concentration in the soil after salt imposition corresponded to 100 mM NaCl (B). Plant responses to salinity stress were monitored for 7 days using PlantScreen TM System by image-based morphometric analysis and in-depth analysis of chlorophyll fluorescence kinetics (C). Materials and Methods RGB imaging, morphometric and color index analysis Results 0 0.2 0.4 0.6 0.8 1 1.2 1.4 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 0 0.2 0.4 0.6 0.8 1 1.2 1.4 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 0 1 2 3 4 5 6 7 Col-0 C24 Fv/Fm Fv´/Fm´ Fq´/Fm´ NPQ qP Fv/Fm Fv´/Fm´ Fq´/Fm´ NPQ qP Photosynthetic performance is rapidly reduced in salt treated plants . Salinity induced rapid changes in regulatory light-induced heat dissipation (NPQ), PSII operating efficiency (Fq´/Fm´), photochemical quenching (qP) and partially in maximum quantum yield in light-adapted state (Fv´/Fm´). No salinity induced changes occured for Fv/Fm parameter. Growth rate in salt treated plants was reduced early upos stress imposition with C24 showing increased salt tolerance Integrative concept of PlantScreen TM high-throughput phenotyping platform provides a powerful tool for acquisition and selection of morphological and physiological parameters. Rapidly after stress initiation photosynthetic performance of the salt-treated plants was compromised, followed by growth retardation and changes in greenness. Presented method shows robust experimental set-up for salinity tolerance screening in Arabidopsis and other plant species. Acknowledgements This work was carried out at PPRC at Photon Systems Instruments (Czech Republic) with partial financial support through IDP Bridges Marie Curie Initial Training Network. Contact: panzarova@ psi.cz Conclusions Control Salt Col-0 C24 Col-0 Days after NaCl treatment Area (mm 2 ) 0 200 400 600 800 1000 0 1 2 3 4 5 6 7 0 200 400 600 800 1000 0 1 2 3 4 5 6 7 Control Salt C24 * * * * Relative changes in rosette color are affected by salt stress treatment . 100% stacked charts of 9 RGB color-coded greenness hues presented as changes in % area over time. The greenness hues summarize the [red:green:blue] channel values corresponding to the green hues identified through the color-segmentation process of RGB images. Salt treated Control Days after NaCl treatment 0 1 3 5 7 0 1 3 5 7 0 1 3 5 7 0 1 3 5 7 Control Salt Control Salt Col-0 C24 Salt treated Control

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Page 1: AUTOMATED PHENOTYPING SYSTEMS Compactnessplantphenotyping.com/documents/poster.pdf · 2018-04-25 · PPRC is situated in countryside next to Brno in Czech Republic, city where Johann

Mariam Awlia1, Arianna Nigro2, Jiří Fajkus3, Martin Trtílek3, Diana Santelia2, Mark A. Tester1, Magdalena M. Julkowska1 and Klára Panzarová3

1King Abdullah University of Science and Technology (KAUST), Thuwal, Saudi Arabia; 2Institute of Plant Biology, University of Zurich, Switzerland; 3PSI (Photon Systems Instruments), Czech Republic

Soil salinity is one of the main stress factors severely affecting theagriculture land in global scale and causing significant reduction of plantgrowth and yield. To enhance our understanding of the early responses tosalinity, we designed an experimental protocol based on automatedintegrative analysis of photosynthetic performance, growth analysis andcolor index analysis at the onset and early phase of salinity stressresponse in Arabidopsis thaliana ecotypes grown in soil. Here we show thatthe developed experimental procedure allows to analyse dynamicallystructural and physiological phenotypes very early upon stress imposition.Results for two accessions Col-0 and C24 are shown. C24 was previouslydescribed for increased salt tolerance. Salinity significantly and rapidlyaffected photosynthetic performance and impacted growth dynamics ofArabidopsis plants at different stages of stress response.

PLANT PHENOTYPING

RESEARCH CENTERScreening tools for identification of traits contributing

to salinity tolerance in Arabidopsis

PLANTSCREENTM

AUTOMATED PHENOTYPING SYSTEMS

www.plantphenotyping.comwww.psi.cz

The mission of the PSI Plant Phenotyping Research

Center (PPRC) is to provide state-of-art infrastructure

for plant cultivation and automated high-throughput

phenotyping of wide range of plant traits.

We offer access to cutting edge instruments and

provide professional support of highly skilled technical

and scientific personnel. PPRC infrastructure is

available for use by visiting scientists and on fee-for-

service basis for a wide range of phenotyping

experiments.

PPRC operates high-end walk-in growth chambers

for precise growth of plants and PlantScreenTM

platforms for automated phenotyping of small and mid-

size plants in controlled environment (e.g. turfgrass,

Arabidopsis thaliana) and for cultivation and

monitoring of larger crop plants up to 1.5 meter in

height.

Chlorophyll fluorescence kinetic imaging and analysis of

photosynthetic performance

Max light adapted fluorescence (Fp)

Time (us)

Time (us)

Time (us)Time (us)

Time (us) Time (us) Time (us)Time (us)

Time (us)Time (us)

Time (us)

Time (us)

Time (us)

Time (us) Time (us)

Time (us)

Time (us)

Time (us)

Time (us)

Time (us)

PAM light LED panel for kinetic imaging

➢ FO, FM, FV, FO', FM', FV', Ft

➢ Max quantum efficinecy Fv/Fm

➢ Photochemical quenching

➢ Non-photochemical quenching

➢ Vitality index

➢ FV'/FM', PhiPSII , qN, qP

Fluorescence kinetics measurement

Mask application and bckgr substraction

Fluorescence quenching kinetics

Original image

Barrel distortion correction

Color segmentation

Background subtraction

Mask detection➢ Area➢ Perimeter

➢ Roundness

➢ Compactness

➢Leaf development tracking

➢ Greening index➢ RLGR

PPRC is situated in countryside next to Brno in Czech

Republic, city where Johann Gregor Mendel lived and

worked. For more information contact us at [email protected].

Automated phenotyping of up to 320 small- and

mid-size scale plants in controlled environment

in PlantScreenTM Compact System.

Automated phenotyping of 270 plants up to 1.5

in height in greenhouse environment in

PlantScreenTM Modular System.

Large-scale walk in chambers for highly precise

plant cultivation.

RationalePhenotyping Protocol

Arabidopsis thaliana Col-0 and C24 accessions were grown in 12h-12h lightconditions under cool-white LED illumination of 150 µmol/m2/s in FS-WIChamber (PSI). Before the salt imposition plants were automaticallyweighed and watered using PlantScreenTM Phenotyping System to adjust soilmoisture to 60% of soil water capacity (A). At 21 days after stratification(DAS) plants were treated with 250 mM NaCl solution for one hour, ensuringsaturation of the soil with the solution. The effective NaCl concentration inthe soil after salt imposition corresponded to 100 mM NaCl (B). Plantresponses to salinity stress were monitored for 7 days using PlantScreenTM

System by image-based morphometric analysis and in-depth analysis ofchlorophyll fluorescence kinetics (C).

Materials and Methods

RGB imaging, morphometric and color index analysis

Results

0

0.2

0.4

0.6

0.8

1

1.2

1.40

1 23

45

67

0

1

2

3

4

5

67

01

234567

01

23

4

5

6

7

0

1

23

45

67

0

0.2

0.4

0.6

0.8

1

1.2

1.40

1 23

45

67

0

1

2

3

4

5

67

01

234567

01

23

4

5

6

7

0

1

23

45

67

Col-0 C24

Fv/Fm

Fv´/Fm´

Fq´/Fm´

NPQ

qP Fv/Fm

Fv´/Fm´

Fq´/Fm´

NPQ

qP

Photosynthetic performance is rapidly reduced in salt treated plants . Salinity induced rapid changes in regulatory light-induced heat dissipation (NPQ), PSII

operating efficiency (Fq´/Fm´), photochemical quenching (qP) and partially in maximum quantum yield

in light-adapted state (Fv´/Fm´). No salinity induced changes occured for Fv/Fm parameter.

Growth rate in salt treated plants was reduced early upos stress imposition with C24 showing increased salt tolerance

Integrative concept of PlantScreenTM high-throughput phenotyping platform provides a powerful tool for acquisition and selection of morphological and

physiological parameters. Rapidly after stress initiation photosynthetic performance of the salt-treated plants was compromised, followed by growth retardation

and changes in greenness. Presented method shows robust experimental set-up for salinity tolerance screening in Arabidopsis and other plant species.

AcknowledgementsThis work was carried out at PPRC at Photon Systems Instruments (Czech Republic)

with partial financial support through IDP Bridges Marie Curie Initial Training Network.

Contact: [email protected]

Conclusions

Control Salt

Col-

0

C24

Col-0

Days after NaCl treatment

Area

(mm

2 )

0

200

400

600

800

1000

0 1 2 3 4 5 6 70

200

400

600

800

1000

0 1 2 3 4 5 6 7

Control Salt

C24

**

**

Relative changes in rosette color are affected by salt stress treatment . 100% stacked charts of 9 RGB color-coded greenness hues presented as changes in

% area over time. The greenness hues summarize the [red:green:blue] channel values corresponding

to the green hues identified through the color-segmentation process of RGB images.

Salt treated

Control

Control Salt

0

25

50

75

100

0 3 6 9 0 3 6 9

Days after salt treatment

Gre

enne

ss h

ues

(% A

rea)

Control Salt

0

25

50

75

100

0 3 6 9 0 3 6 9

Days after salt treatment

Gre

enne

ss h

ues

(% A

rea)

Days after NaCl treatment0 1 3 5 7 0 1 3 5 7 0 1 3 5 7 0 1 3 5 7

Control Salt Control Salt

Col-0 C24

Salt treated

Control