Remote sensing – Beyond images Mexico 14-15 December 2013 The workshop was organized by CIMMYT Global Conservation Agriculture Program (GCAP) and funded by the Bill & Melinda Gates Foundation (BMGF), the Mexican Secretariat of Agriculture, Livestock, Rural Development, Fisheries and Food (SAGARPA), the International Maize and Wheat Improvement Center (CIMMYT), CGIAR Research Program on Maize, the Cereal System Initiative for South Asia (CSISA) and the Sustainable Modernization of the Traditional Agriculture (MasAgro)

1. High-resolution phenotyping
Water flow dynamics
Chloroplast movement
Hae Koo Kim, Joonghyuk Park, Jeong Eun Ryu,
Sungsook Ahn, Ildoo Hwang and Sang Joon Lee

2. Plants in Environmental & Energy Sciences
 Diversity in vascular system: Model plants
 Central Role in the Water Cycle on Earth (conifers): water transport over 100m
-Gymnosperms
 Green revolution yet to come
-Angiosperms (monocots & dicots): few cm to tens of m
for smallholder farmers?
 Importance of the Plants
 Agriculture: up to 70% of the
Water & mineral transport : Xylem
water use
 “Blue revolution” in
Agriculture:
Drought stress in water-limited
environments
- Drop by Drop
- Gene by Gene
Pennisi (Science, 2008)
 Water transport in plants
Cohesion-Tension Theory, Böhm, 1893;
Dixon and Joly, 1895)
Scholander et al. (1965) Science 148
Johnson & Dixon (1965) Nature 208
Scholander et al.,1965
Balling & Zimmermann, 1990
Pockman et al. (1995) Nature 378
Holbrook et al. (1995) Science 270

3. Visualization of Water flow at different level on Earth
6,650 km

4. Synchrotron
facilities
PAL, Pohang
PF, Tsukuba
SSRF, Shanghai

5. Visualization of water flow dynamics using synchrotron X-ray imaging
Synchrotron X-ray imaging
-Absorption and phase-contrast based imaging
-High temporal (ms) and spatial resolution (μm)
-Ideal to visualize xylem vessels of monocots
-Real-time dynamics of refilling process
Kim & Lee, 2010 New Phytologist

6. Phenotyping Xylem Structure/Function relationships
Sap flow dynamics in xylem vessels
Xylem
Safety
Efficiency
Structure
Fibers
Vascular bundle
[WATER TRANSPORT]
Efficiency vs. Safety trade-off in water transport
Function
High resistance
Modularity
Support
Repair
Parenchyma
Vessels
Storage
Transport
Low resistance
Connectivity
Phloem
FUNCTION of Xylem: Efficiency & Safety of water transport
STRUCTURAL characteristics of XYLEM
 Sap flow dynamics

7. Efficiency and Safety trade-offs in water conduits
Number of pits
16-20
11-15
6-10
1-5
0
Cluster 1
Cluster 2-2
Cluster 2-1
U
R
8
2
P,
Q
R
4
Pc
P
Cluster 3
8
Metaxylem vessel number and lumen area
• Diameter
A:B:C=4:2 :1
• Vessel area A : B : C = 16 : 4 : 1
• Flow rates A : B : C = 256 : 16 : 1
Cluster
Number (ea)
AVG (µm2)
MAX (µm2)
MIN (µm2)
1
73
226.7 (±102.3)
496.7
66.2
2
83
201.8 (±87.4)
489.6
76.0
3
126
108.7 (±37.5)
318.3
52.9
7

8. Visualization of internal structure of plants and flow dynamics
Kim and Lee., 2010 New Phytologist
Ahn et al., 2010 ACS Nano

9. -Long distance water transport (efficiency)
-Short distance, higher resistance, local water distribution (safety)

10. Synchrotron X-ray CT: Vascular bundle organization in maize leaf

11. 3D organization of Vascular bundles in maize leaf

12. Xylem anatomical characteristics
• Safety: Regulation of -Axial flow dynamics by perforation plates
-Radial flow dynamics by pit membranes & network connexion
Basis of embolism,
cavitation
mechanism
Environmental
effects:
transpiration, osm
ostic stress
• Safety feature
of perforation
plates, lateral
pits: air
bubbles
appearance/re
moval, direct
radial inflow of
water
 Network
• Efficiency:
-Effect of enhanced
transpiration
-Long distance water
transport system
In planta sap
flow tracking
Protoxylem (PX)
Metaxylem (MX)
•Use of various type of particles to investigate xylem cell
wall properties (interactions of AuNPs with cell walls)

13. Flow dynamics in dicot plants (Arabidopsis) of dicot xylem structure
Arabidopsis as model
Experimental method:
-Synchrotron X-ray CT
[Xylem network 3D organization]
-Hydrophilic AuNP solution
[Indicator of sap flow rate]
AuNP-OH
Xylem SCW
HO
CH2
CH2
CH2
CH2
OH
Difference of xylem
vessel activity monitored
by AuNP staining profile
13

16. Pattern of xylem vessel activity
in normal condition
Normal uptake profile
(transpiration driven)
Pattern of xylem vessel activity
generated by external pressure
Artificial uptake profile
(external pressure driven)

17. Visualization of water transport pathways
Hydrophilic gold-nanoparticles (AuNP) :
tracers of axial & radial water flow pathways
c
Interestingly, all metaxylem vessels do not show the same level of
activity for water transport
17

18. Regulation of water transport along plant height
Differences in vascular bundle organization and pressure gradient
directly impact AuNPs staining profile along the height of the
inflorescence stem
18

19. Water flow pathway in the whole plant scale
Efficient and safe water flow may coexist with one system as
division into long-distance and local distribution
19

20. Visualization of chloroplast movement using two-photon microscopy
Two-photon microscopy
Tube
lens
Scan
lens
Ti:Sapphire
laser
Dichroic mirror
DM1
Tube Scan
lens lens
Dichroic mirror
DM2
Galvanometer
scanner
PMT
Piezo stage
Filter
Objective
lens
Computer
Translation stage

21. Monitoring of chloroplast movement depending on light
S
S
S
M
BS
S
S
S

22. Summary
Place of plants in Environmental & Energy Sciences
Importance of understanding Plant-Water relation
Synchrotron X-ray imaging method is a powerful
tool to study plant vascular structure and sap flow
dynamics
-Monocot and dicot plants 3D xylem structure
(“Wiring diagram” of plant microfluidic system)
-Demand-driven flow system
-Efficiency and safety of water transport
Perspectives of High-resolution phenotyping
-Nano-CT of xylem
-Optical Coherance tomography
-Two-photon microscopy (photosynthesis)

23. Acknowledgements
Advanced Biomass R&D Center
Prof. Sang Joon Lee (Department of
Mechanical Engineering, POSTECH)
Prof. Il Doo Hwang (Department of
Life Sciences, POSTECH)
Dr. Sung Sook Ahn
Joong Hyuk Park
Jeong Eun Ryu
Synchrotron
facilities
PAL, Pohang
SSRF, Shanghai
PF, Tsukuba