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M.Sc course seminar On the topic
Supervisor : Prof. J. P Shahi Name : Kumari ShikhaI.D. NO. G-15096 M.Sc (Ag) ,final year
Department of Genetics and plant breedingBanasars Hindu University
Varanasi-2210055/24/2017 1
CONTENTS Introduction
Importance
Imaging technology
Gloss of modern technologies
Traits phenotyped
Relevance
Data management
conclusion
3-D ImagingFar –infrared imagingNIR ImagingFluorescence imagingAutomated watering and sprayingHyperspectral imagingVisible imaging
5/24/2017 2
Plant High-throughput Phenotyping
• Phenomics term given by Gerlai, 2002.
• The term phenomic refers to sum total of phenotypes
at various levels ranging from molecules to organs
and the whole organism.
• Study of plant growth ,architecture, performance and
composition using high throughput methods of data
acquisition and analysis.
5/24/20173
Continue..
• Phenotyping is essential for –
functional analysis of specific genes
forward and reverse genetic analysis
production of elite plants
• High throughput is essential for phenotyping in
different growth conditions of many different lines.
mutant populations
mapping population
breeding population
germplasm collection 5/24/2017 4
Forward phenomics vs Reverse phenomics
5/24/2017 5
Why phenotyping needed?
Need more food
By 2050, 9.1 billion people Efficient breeding technique required
Tilman, et al5/24/2017 6
Accelerating Plant Phenomics
High-throughput screens
Multiple camera units
Non-destructive measurements
Quantitative analysis
Monitor gowth dynamics
Stress assessment
Link to genomics
Opening new prospects
5/24/2017 7
Automated Facilities for HTP
Australian plant phenotypingnetworking
ICAR-IIHR5/24/2017 8
Imaging Technique
3-D imaging
Near infrared imaging
Far infrared imaging
Flourescence imaging
Visible light scanning
Automated spraying and watering
High throughput Plant Phenotyping1486203666711
5/24/2017 9
3-Dimensional imaging
SIDE VIEW SIDE VIEW TOP VIEW
After 90 degree rotation
Digital photos of the top and side view of a plant combined into 3D image.
5/24/201710
Measurements taken using 3-D
• Shoot mass
• Leaf number, shape and angle
• Leaf colour
• Leaf health
5/24/2017 11
Far-infrared Imaging
• Measure temperature difference between leaves and plants.
• Temperature ranges between 15-1000 micrometer.
• Cooler plants absorb more water
• Can be used for single plant to whole plant
• Temperature differences used to measure:-
Photosynthetic activity
Salinity tolerance
Effective water use efficiency
5/24/2017 12
Near- infrared imaging
• More the presence of chlorophyll more will be
reflectance in NIR range.
• Facilitates estimation of
Water content and movement within leaves and soil
Carbohydrate content of leaves
5/24/2017 13
Spongy layer present backside of leaves that reflects lot of light in NIR range,after dehydration or stress condition this spongy layer get collapsed hencereflect less NIR light but same visible range. Thus , we can differentiate healthyplant from sickly plant.
5/24/2017 14
Fluorescence imaging
Measure photosynthesis ratebiotic and abiotic stress responsesChlorophyll content.
5/24/2017 15
Visible light scanning (VIS)
• Colour information give estimation of the degreeof senescence.
• Senescence of older leaves during drought suggestan escape or avoidance.
• Genotypes with stay-green type can be identifiedthat would be able to continue photosynthesisunder water stress
5/24/2017 16
Continued..
Measure aspects of plant architecture such as
• Image-based projected biomass, leaf area, colour, growth dynamics, seedling vigour, seed morphology,
• Root architecture, leaf disease severity assessments, yield, and fruit number and distribution.
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Leaf growth decreases in response to drought even before a decline in stomatal conductance or photosynthesis.
5/24/2017 18
Hyperspectral Imaging
• Spectral reflectance is the fraction of light reflected
by non transparent surface.
• Researchers use this spectral reflectance fordetection of plants stressed by saline soil ordrought, well before it can be eye.
5/24/2017 19
Positron emission tomography(PET)
• Nuclear imaging system
• Produces 3-D image of a functional process
• While consumption of co2 , transport of
radiolabelled carbon imaged in 3-D by PET.
5/24/2017 20
Root Phenotyping
• Different procedure-
Visualization of excavated root system.
Analysis by camera systems which are introduced into
soil through small tubes made up of Plexiglass (changes in
electrical properties of soil due to water uptake by soil).
2-D and 3-D analysis
Phenotyping platforms using aeroponic or hydroponic
culture systems for direct visualization and imaging of roots.
5/24/2017 21
Steps used in the analysis ofroot morphology.Step 1: soil coring;
Step 2: dividing soil samples
Step 3: washing of roots
Step 4: separating roots fromsoil
Step 5: preserving roots in 25%ethanolStep 6: scanning roots using rootscanner Epson Perfection
step 7 analyzing the scannedroot images using Winrhizosoftware.
2-D Phenotyping of root
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3-D PHENOTYPING
5/24/2017 23
Rhizoponics is an innovative design of hydroponic rhizotronsadapted to Arabidopsis thaliana. The setup allows tosimultaneously characterize the RSA and shoot development fromseedling to adult stages, i.e. from seed to seed. This system offersthe advantages of hydroponics such as control of rootenvironment and easy access to the roots for measurements orsampling. Being completely movable and low cost, it can be usedin controlled cabinets.
Hydroponic
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Measurement of leaf area
• Morphometric method
• Optical flow method
• Particle /marker tracking method
Plant biomass estimation
• 3-digital imaging technique
• Hyperspectral imaging
• Non-optical method (electrical determination of water content of plant , portable nuclear magnetic resonance device)
Seed and fruit phenotyping
• 3-D Laser –scanning technology
• Visual imaging
• NIR Spectroscopy5/24/2017 25
Analysis of root system
• Rootreader 2D
• Smartreader
• Rootreader 3D
Analysis of shoot system
• Hyperspectral imaging
• Visible imaging
Analysis of chemical content
• Mass spectrometry and Gas chromatography (amino acids present in fresh plant material )
• Liquid chromatography
• Flow cytometry
• NIR Spectroscopy5/24/2017 26
Analysis of physiological parameters
• SPAD chlorophyll meter
• Fluorescent imaging (chlorophyll fluorescence show negative co-relation with photosynthetic activity)
Assessment of water use
• CID (carbon isotope discrimination technique used in wheat)
• Leaf and canopy temperature ( higher in case of decreased transpiration rate)
• SPAD chlorophyll meter
Assessment of soil water content
• Mobile NIR
• Visual spectrophotometer
5/24/2017 27
Relevance of HTP
• Identification of stress.
• Rapid and efficient screening for mutants.
• Detection and monitoring of disease epidemics in field.
• Detection of root attack by pathogens.
• Facilitate screening of germplasm.
• Study of various physiological processes.
• Modelling of biomass production.
5/24/2017 28
continued..
Facilitate selection of superior genotypes from breeding population.
Allow huge genomic information to be reliably related to specific phenotypes.
Permit systematic study of pleiotropic effects of the genes.
Crop improvement .
5/24/2017 29
Data management
Phenomic data management involves three critical components:
ALGORITHM AND PROGRAM
PHENOTYPIC INFORMATION
SENSORY DATA
MODEL DEVELOPMENT
GENOTYPE AND PHENOTYPE
INTERACTIONSUNDERSTAND
MANAGEMENT DATABASES
RESOURCE DEVELOPMENT AND RESOURCE SHARING
5/24/2017 30
Continued
Some suggestion :-
Creating complete and accurate metadata
Deposition of data into primary repository
Easily Accessible to every researchers
Development of open source community database
5/24/2017 31
Software companies
• Some of these companies encourage co-development (system customization, softwaredevelopment for computation) as a process ofimproving their current product and productutilization such as Lemnashare and Lemnalauncher etc.
PHENOSPEX5/24/2017 32
Case study
• GiNA platform developed for phenotyping of horticultural crops. For small fruits such as grapes, cranberries, or cherries a picture of 40 fruits can be taken every minute (or less). Therefore, in an hour, at least 20 different parameters for 2400 fruits can be accurately measured from 60 images.
LENGTH , WIDTH , PERIMETER AREA , COLOR ESTIMATED
POTATO AND CHERRY FRUIT5/24/2017 33
Access genotypic adaptation to water stress
Thermography
Leaf temperature detection byinfrared thermometers hasbeen used to detect waterstress, which results instomatal closure and anincrease in temperaturethrough decreased adiabaticcooling
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a): Excavated maize root stocks which were split lengthwiseb): imaged root stock under constant illuminationc): root top angled): gap size distribution e): cluster thickness distribution Colombi et al.2015
5/24/2017 35
This high-throughput phenotypingsystem developed at the USDA Arid-Land Agricultural Research Center inMaricopa, AZ is being used to collectplant height, canopy temperature, andcanopy reflectance data from cottonplants.
Researchers at CSIRO using aremote-controlled gas-poweredmodel helicopter called the“phenocopter” to measure plantheight, canopy cover, lodging, andtemperature throughout a day.
5/24/2017 36
Future of HTP
• Current phenotyping is largely extensive hence need for intensive approach.
• Available methods are not satisfactory , hence urgent need to develop suitable statistical models.
• Software developed for statistical analysis should permit automated data analysis.
• Integration of all phenomic related research as did for genomic efforts.
• Phenomics teams must be transdisciplinary.
Bilder et al. 2009
5/24/2017 37
Phenotyping
Conclusion
Identifying differences among genotypes andinterpretation of detected differences.
(Fiorani and Schurr, 2013; Klukaset al., 2014)
Analytical approach
Management
5/24/2017 38
Thankyou5/24/2017 39