Reproducible and efficient high-throughput phenotyping approaches, combined with advances in genome sequencing, are facilitating the discovery of genes affecting plant performance. Salinity tolerance is a desirable trait that can be achieved through breeding, where most have aimed at selecting for plants that perform effective ion exclusion from the shoots. To determine overall plant performance under salt stress, it is helpful to investigate several plant traits collectively in one experimental setup. Hence, we developed a quantitative phenotyping protocol using a high-throughput phenotyping system, with RGB and chlorophyll fluorescence (ChlF) imaging, which captures the growth, morphology, color and photosynthetic performance of Arabidopsis thaliana plants in response to salt stress. We optimized our salt treatment by controlling the soil-water content prior to introducing salt stress. We investigated these traits over time in two accessions in soil at 150, 100, or 50 mM NaCl to find that the plants subjected to 100 mM NaCl showed the most prominent responses in the absence of symptoms of severe stress. In these plants, salt stress induced significant changes in rosette area and morphology, but less prominent changes in rosette coloring and photosystem II efficiency. Clustering of ChlF traits with plant growth of nine accessions maintained at 100 mM NaCl revealed that in the early stage of salt stress, salinity tolerance correlated with non-photochemical quenching processes and during the later stage, plant performance correlated with quantum yield. This integrative approach allows the simultaneous analysis of several phenotypic traits. In combination with various genetic resources, the phenotyping protocol described here is expected to increase our understanding of plant performance and stress responses, ultimately identifying genes that improve plant performance in salt stress conditions.

Division of Biological and Environmental Sciences and Engineering King Abdullah University of Science and Technology Thuwal Saudi Arabia

Institute of Plant and Microbial Biology University of Zurich Zurich Switzerland

PSI Drásov Czech Republic

Zobrazit více v PubMed

Baker N. R. (2008). Chlorophyll fluorescence: a probe of photosynthesis in vivo. Annu. Rev. Plant Biol. 59 89–113. 10.1146/annurev.arplant.59.032607.092759 PubMed DOI

Baker N. R., Rosenqvist E. (2004). Applications of chlorophyll fluorescence can improve crop production strategies: an examination of future possibilities. J. Exp. Bot. 55 1607–1621. 10.1093/jxb/erh196 PubMed DOI

Ben Abdallah S., Aung B., Amyot L., Lalin I., Lachaal M., Karray-Bouraoui N., et al. (2016). Salt stress (NaCl) affects plant growth and branch pathways of carotenoid and flavonoid biosyntheses in Solanum nigrum. Acta Physiol. Plant. 38:72 10.1007/s11738-016-2096-8 DOI

Berger B., de Regt B., Tester M. (2012). Trait dissection of salinity tolerance with plant phenomics. Methods Mol. Biol. 913 399–413. 10.1007/978-1-61779-986-0_27 PubMed DOI

Bresson J., Vasseur F., Dauzat M., Koch G., Granier C., Vile D. (2015). Quantifying spatial heterogeneity of chlorophyll fluorescence during plant growth and in response to water stress. Plant Methods 11:23 10.1186/s13007-015-0067-5 PubMed DOI PMC

Brestic M., Zivcak M. (2013). “PSII fluorescence techniques for measurement of drought and high temperature stress signal in crop plants: protocols and applications,” in Molecular Stress Physiology of Plants, eds Rout R. G., Das B. A. (New Delhi: Springer; ), 87–131.

Bridge L. J., Franklin K. A., Homer M. E. (2013). Impact of plant shoot architecture on leaf cooling: a coupled heat and mass transfer model. J. R. Soc. Interface 10:20130326 10.1098/rsif.2013.0326 PubMed DOI PMC

Brown T. B., Cheng R., Sirault X. R., Rungrat T., Murray K. D., Trtilek M., et al. (2014). Traitcapture: genomic and environment modelling of plant phenomic data. Curr. Opin. Plant Biol. 18 73–79. 10.1016/j.pbi.2014.02.002 PubMed DOI

Cabot C., Sibole J. V., Barcelo J., Poschenrieder C. (2014). Lessons from crop plants struggling with salinity. Plant Sci. 226 2–13. 10.1016/j.plantsci.2014.04.013 PubMed DOI

Campbell M. T., Knecht A. C., Berger B., Brien C. J., Wang D., Walia H. (2015). Integrating image-based phenomics and association analysis to dissect the genetic architecture of temporal salinity responses in rice. Plant Physiol. 168 1476–1489. 10.1104/pp.15.00450 PubMed DOI PMC

Chaves M. M., Flexas J., Pinheiro C. (2009). Photosynthesis under drought and salt stress: regulation mechanisms from whole plant to cell. Ann. Bot. 103 551–560. 10.1093/Aob/Mcn125 PubMed DOI PMC

Chen T. W., Kahlen K., Stutzel H. (2015). Disentangling the contributions of osmotic and ionic effects of salinity on stomatal, mesophyll, biochemical and light limitations to photosynthesis. Plant Cell Environ. 38 1528–1542. 10.1111/pce.12504 PubMed DOI

Dhondt S., Wuyts N., Inze D. (2013). Cell to whole-plant phenotyping: the best is yet to come. Trends Plant Sci. 18 433–444. 10.1016/J.Tplants.2013.04.008 PubMed DOI

Fricke W., Akhiyarova G., Wei W. X., Alexandersson E., Miller A., Kjellbom P. O., et al. (2006). The short-term growth response to salt of the developing barley leaf. J. Exp. Bot. 57 1079–1095. 10.1093/Jxb/Erj095 PubMed DOI

Ghanem M. E., Marrou H., Sinclair T. R. (2015). Physiological phenotyping of plants for crop improvement. Trends Plant Sci. 20 139–144. 10.1016/j.tplants.2014.11.006 PubMed DOI

Godfray H. C., Beddington J. R., Crute I. R., Haddad L., Lawrence D., Muir J. F., et al. (2010). Food security: the challenge of feeding 9 billion people. Science 327 812–818. 10.1126/science.1185383 PubMed DOI

Hairmansis A., Berger B., Tester M., Roy S. J. (2014). Image-based phenotyping for non-destructive screening of different salinity tolerance traits in rice. Rice 7:16 10.1186/s12284-014-0016-3 PubMed DOI PMC

Hannah M. A., Wiese D., Freund S., Fiehn O., Heyer A. G., Hincha D. K. (2006). Natural genetic variation of freezing tolerance in Arabidopsis. Plant Physiol. 142 98–112. 10.1104/pp.106.081141 PubMed DOI PMC

Henley W. J. (1993). Measurement and interpretation of photosynthetic light-response curves in algae in the context of photoinhibition and diel changes. J. Phycol. 29 729–739. 10.1111/j.0022-3646.1993.00729.x DOI

Humplik J. F., Lazar D., Husickova A., Spichal L. (2015). Automated phenotyping of plant shoots using imaging methods for analysis of plant stress responses - a review. Plant Methods 11:29 10.1186/s13007-015-0072-8 PubMed DOI PMC

James R. A., Munns R., Von Caemmerer S., Trejo C., Miller C., Condon T. (2006). Photosynthetic capacity is related to the cellular and subcellular partitioning of Na, K and Cl in salt-affected barley and durum wheat. Plant Cell and Environment 29 2185–2197. 10.1111/J.1365-3040.2006.01592.X PubMed DOI

Jansen M., Gilmer F., Biskup B., Nagel K. A., Rascher U., Fischbach A., et al. (2009). Simultaneous phenotyping of leaf growth and chlorophyll fluorescence via GROWSCREEN FLUORO allows detection of stress tolerance in Arabidopsis thaliana and other rosette plants. Funct. Plant Biol. 36 902–914. 10.1071/Fp09095 PubMed DOI

Junker A., Muraya M. M., Weigelt-Fischer K., Arana-Ceballos F., Klukas C., Melchinger A. E., et al. (2015). Optimizing experimental procedures for quantitative evaluation of crop plant performance in high throughput phenotyping systems. Front. Plant Sci. 5:770 10.3389/fpls.2014.00770 PubMed DOI PMC

Lazar D. (2015). Parameters of photosynthetic energy partitioning. J. Plant Physiol. 175 131–147. 10.1016/j.jplph.2014.10.021 PubMed DOI

Longenberger P. S., Smith C. W., Duke S. E., McMichael B. L. (2009). Evaluation of chlorophyll fluorescence as a tool for the identification of drought tolerance in upland cotton. Euphytica 166 25–33. 10.1007/s10681-008-9820-4 DOI

Ma S. S., Gong Q. Q., Bohnert H. J. (2006). Dissecting salt stress pathways. J. Exp. Bot. 57 1097–1107. 10.1093/Jxb/Erj098 PubMed DOI

Maxwell K., Johnson G. N. (2000). Chlorophyll fluorescence - a practical guide. J. Exp. Bot. 51 659–668. 10.1093/jexbot/51.345.659 PubMed DOI

Mishra K. B., Iannacone R., Petrozza A., Mishra A., Armentano N., La Vecchia G., et al. (2012). Engineered drought tolerance in tomato plants is reflected in chlorophyll fluorescence emission. Plant Sci. 182 79–86. 10.1016/j.plantsci.2011.03.022 PubMed DOI

Munns R., James R. A., Sirault X. R., Furbank R. T., Jones H. G. (2010). New phenotyping methods for screening wheat and barley for beneficial responses to water deficit. J. Exp. Bot. 61 3499–3507. 10.1093/jxb/erq199 PubMed DOI

Munns R., Tester M. (2008). Mechanisms of salinity tolerance. Annu. Rev. Plant Biol. 59 651–681. 10.1146/annurev.arplant.59.032607.092911 PubMed DOI

Muranaka S., Shimizu K., Kato M. (2002). A salt-tolerant cultivar of wheat maintains photosynthetic activity by suppressing sodium uptake. Photosynthetica 40 509–515. 10.1023/A:1024335515473 DOI

Murchie E. H., Lawson T. (2013). Chlorophyll fluorescence analysis: a guide to good practice and understanding some new applications. J. Exp. Bot. 64 3983–3998. 10.1093/jxb/ert208 PubMed DOI

Oxborough K. (2004). Imaging of chlorophyll a fluorescence: theoretical and practical aspects of an emerging technique for the monitoring of photosynthetic performance. J. Exp. Bot. 55 1195–1205. 10.1093/jxb/erh145 PubMed DOI

Oxborough K., Baker N. R. (1997). Resolving chlorophyll a fluorescence images of photosynthetic efficiency into photochemical and non-photochemical components - calculation of qP and Fv’/Fm’ without measuring Fo’. Photosynth. Res. 54 135–142. 10.1023/A:1005936823310 DOI

Rajendran K., Tester M., Roy S. J. (2009). Quantifying the three main components of salinity tolerance in cereals. Plant Cell Environ. 32 237–249. 10.1111/J.1365-3040.2008.01916.X PubMed DOI

Rascher U., Liebig M., Luttge U. (2000). Evaluation of instant light-response curves of chlorophyll fluorescence parameters obtained with a portable chlorophyll fluorometer on site in the field. Plant Cell and Environ. 23 1397–1405. 10.1046/j.1365-3040.2000.00650.x DOI

Roy S. J., Negrao S., Tester M. (2014). Salt resistant crop plants. Curr. Opin. Biotechnol. 26 115–124. 10.1016/j.copbio.2013.12.004 PubMed DOI

Sirault X. R. R., James R. A., Furbank R. T. (2009). A new screening method for osmotic component of salinity tolerance in cereals using infrared thermography. Funct. Plant Biol. 36 970–977. 10.1071/Fp09182 PubMed DOI

Stephan A. B., Schroeder J. I. (2014). Plant salt stress status is transmitted systemically via propagating calcium waves. Proc. Natl. Acad. Sci. U.S.A. 111 6126–6127. 10.1073/Pnas.1404895111 PubMed DOI PMC

Stepien P., Johnson G. N. (2009). Contrasting responses of photosynthesis to salt stress in the glycophyte Arabidopsis and the halophyte Thellungiella: role of the plastid terminal oxidase as an alternative electron sink. Plant Physiol. 149 1154–1165. 10.1104/Pp.108.132407 PubMed DOI PMC

Tester M., Langridge P. (2010). Breeding technologies to increase crop production in a changing world. Science 327 818–822. 10.1126/science.1183700 PubMed DOI

Tilman D., Balzer C., Hill J., Befort B. L. (2011). Global food demand and the sustainable intensification of agriculture. Proc. Natl. Acad. Sci. U.S.A. 108 20260–20264. 10.1073/Pnas.1116437108 PubMed DOI PMC

Van Oosten M. J., Sharkhuu A., Batelli G., Bressan R. A., Maggio A. (2013). The Arabidopsis thaliana mutant air1 implicates SOS3 in the regulation of anthocyanins under salt stress. Plant Mol. Biol. 83 405–415. 10.1007/s11103-013-0099-z PubMed DOI

Zhang X., Hause R. J., Jr., Borevitz J. O. (2012). Natural genetic variation for growth and development revealed by high-throughput phenotyping in Arabidopsis thaliana. G3 (Bethesda) 2 29–34. 10.1534/g3.111.001487 PubMed DOI PMC

Differential expression and localization of expansins in Arabidopsis shoots: implications for cell wall dynamics and drought tolerance

Genotyping-by-sequencing uncovers a Thinopyrum 4StS·1JvsS Robertsonian translocation linked to multiple stress tolerances in bread wheat

Integrative phenotyping analyses reveal the relevance of the phyB-PIF4 pathway in Arabidopsis thaliana reproductive organs at high ambient temperature

The Absence of the AtSYT1 Function Elevates the Adverse Effect of Salt Stress on Photosynthesis in Arabidopsis

Crucial Cell Signaling Compounds Crosstalk and Integrative Multi-Omics Techniques for Salinity Stress Tolerance in Plants

Seed Priming With Protein Hydrolysates Improves Arabidopsis Growth and Stress Tolerance to Abiotic Stresses

Integration of Phenomics and Metabolomics Datasets Reveals Different Mode of Action of Biostimulants Based on Protein Hydrolysates in Lactuca sativa L. and Solanum lycopersicum L. Under Salinity

Glucose uptake to guard cells via STP transporters provides carbon sources for stomatal opening and plant growth

A Novel Image-Based Screening Method to Study Water-Deficit Response and Recovery of Barley Populations Using Canopy Dynamics Phenotyping and Simple Metabolite Profiling

A Combined Phenotypic and Metabolomic Approach for Elucidating the Biostimulant Action of a Plant-Derived Protein Hydrolysate on Tomato Grown Under Limited Water Availability

Understanding the Biostimulant Action of Vegetal-Derived Protein Hydrolysates by High-Throughput Plant Phenotyping and Metabolomics: A Case Study on Tomato

An Automated Method for High-Throughput Screening of Arabidopsis Rosette Growth in Multi-Well Plates and Its Validation in Stress Conditions