1. GCP 2011General Research
Meeting
21-25 Sep 2011
L Krishnamurthy
ICRISAT
Chickpeas - Phenotyping for drought traits

2. • Trait’s relevance for phenotyping
• Framework for trait choice: Analytical
model explaining yield
• Phenotyping experience for
- Root structure and function (TL1)
- Canopy temperature
- TE through Δ13C (GCP : SP3 G4008-12)
- Rate of partitioning
- For drought tolerance indices
Chickpeas - Phenotyping for drought traits

3. Drought type - strategies (intensity/timing/duration)
Trait that matters & linked to yield
Link trait-environment
Constitutive vs adaptive traits
Non-stress related factors affecting yield
e.g.: Time to flowering & Yield potential
Expl. design – Data quality
Simple, precise, high throughput amenable
Choice of traits to phenotype

4. • Phenotyping for root system
Structural and Functional
J. Kashiwagi, L. Krishnamurthy, H.D. Upadhyaya,
P.M. Gaur, R.K. Varshney,
Z.A. Mainassara, V. Vadez and others

5. •Establishing the contribution of root traits to DRT yield in
field,
•Assessment of root variation in field using an RILs
•Establishment of acceptable protocols that produce closely
related root performance in cylinders
•Phenotypic assessment for root related traits in the
minicore germplasm, three mapping populations and the
reference collection
•And identification of robust QTLs are the achievements of
the past decade
Chickpeas - Phenotyping for root traits

7. Variation in rooting depthRootingdepth(cm)
85
90
95
100
105
110
115
120
125
130
Mini-core germplasm accessions (n=211) plus 5 popular cultivars grown in cylinder systems and
sampled at 35-d after sowing.
ICC8261
ICC
4958
Annigeri
ICC 283
ICC
1882
SEd=12.95

8. Variation in root length density
Mini-core germplasm accessions (n=211) plus 5 popular cultivars grown in cylinder systems and
sampled at 35-d after sowing.
0.18
0.20
0.22
0.24
0.26
0.28
0.30
0.32Rootlengthdensity(cmcm-3)
ICC8261
ICC4958
Annigeri
ICC283
ICC1882
SEd=0.022

9. ICCM0249
TAA170
GA24
STMS11
CID traits
Root traits
ICCM0249
TAA170
GA24
STMS11
Root + CID + Yield traits
Hot spot for QTL –LG 5
(ICC 4958 × ICC 1882 map)

10. Root function – Transpiration measurements
Dynamic obs.
Root function
i.e. actual transpiration

11. Root phenotyping
Are there simple solutions?

12. Root traits – How useful is the seedling expression?
Contrasting selections of 10 day-old chickpea seedlings from the
reference collection - sand culture.

13. - for a high throughput assessment of
canopy temperature or root function
L. Krishnamurthy, Z.A. Mainassara,
J. Kashiwagi, R. Purushothaman &
V.Vadez
Infrared thermography

14. IR Photograph IR Photo devoid of soil radiation
Advantage over
IR Thermometer:
Observer bias
gets minimized
Canopy temperature
• Simple
• Truly high throughput
• Yet to be established
• Critical stage of
observation

15. 20
22
24
26
28
30
32
34
36
ICC 1882 ICCV10 ICC 867 Annigeri ICC
14799
ICC 283 ICC 8261 ICC
14778
ICC 4958 ICC 3325 ICC 3776 ICC 7184
Optimally irrigated
Drought stressed
Variation in canopy temperature across 8 drought tolerant
and 4 sensitive genotypes at early-pod fill stage, 2009-10.

16. Variation in canopy temperature across 60 drought tolerant
and 24 sensitive genotypes at 75 DAS, 2009-10.
27
28
29
30
31
32
33
34
35
1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76
Germplasm accessions
C.areaxC.temp.(0
C)
25
27
29
31
33
35
37
39
1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76
Canopytemp.(0
C)
0.8
0.9
1
1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76
Canopyproportion
Evap. 5.1mm
Max T. 30.0 0C
Min T. 9.0 0C

17. Well-watered conditions Water stress
Leaf conductance
(g h-1
cm-2
)
Canopy
TºC
Shoot DW Leaf
expansion
Total
Tr
FTSW
Threshold
Shoot DW
42 DAS
Outdoor
42 DAS
GC*
42 DAS 42
DAS
56
DAS
42-56
DAS
56 DAS 56 DAS
ICC867 Low Low Hot Low Low Low High Low
ICC3325 Low Low Hot Low Low High Low
ICC3776 High Cool High Low High
ICC4814 High High Cool Low High Low High
ICC7184 Cool High High High Low
ICC8058 High High Cool High High High High Low High
ICC14778 Low Low Low Low High Low
ICC14799 Low Low Hot Low Low Low Low High Low
1

18. Phenotyping for TE (through Δ13C)

19. How does TE contribute
to drought tolerance?
Yield = Shoot biomass x Harvest index
Transpiration Transpiration Efficiency (TE)=13Cx
Leaf biomass + Stem biomass
LAxLeaf biomass/Leaf area ratio (inverse of specific leaf area: SLA)
SPAD
Increasing SLA=reducing the water loss via leaf area reduction + maintain photosynthesis via
leaf thickness increase

21. … and the optimally irrigated section of the DT field evaluation
of the reference collection of chickpea germplasm

22. Means (BLUPs) and range of various traits observed both under drought stressed
and optimally irrigated environments of the 280 accessions of the reference
collection of chickpea germplasm during 2008-09 postrainy season in a Vertisol. (All
the traits showed significantly different many-fold variation)
___________________________________________________________________
Drought stressed Optimally irrigated
__________________ ___________________
Characteristics Mean Range Mean Range
____________________________________________________________________
Days to 50% flowering 51 39-71 54 40-69
Days to maturity 98 83-120* 115 109-126*
Shoot biomass (kg ha-1
) 3444 2259-4726 5263 3602-6977
Seed yield (kg ha-1
) 1557 510-2250 2057 1004-3029
Harvest index 46 11-61 39.4 17.3-52.8
Pod number m-2
1358 512-2332 1965 802-4553
Seed number m-2
1614 504-3131 2516 831-4553
Seeds pod-1
1.2 1.0-1.5 1.3 1.0-1.9
100 seed weight (g) 16.3 8.8-36.2 15.1 7.2-44.6
Shoot biomass (kg ha-1
day-1
) 35.1 20.7-46.9 45.7 30.6-58.9
Seed yield (kg ha-1
day-1
) 32.2 8.5-48.2 39.9 17.7-50.1
___________________________________________________________________

23. 10 leaves
At 63 DAS, Recently formed
Fully expanded
Primary branch-born
from 10 different plants
Dried at 60°C for three days
C isotope measurements at JIRCAS

24. The relationship between per day productivity (grain yield kg ha-1 day-1) or
WUE with carbon isotope discrimination - 2008-09.
y = -3.6x - 62.9
r2
= 0.11***
0
10
20
30
40
50
60
-28.5 -28.0 -27.5 -27.0 -26.5 -26.0 -25.5 -25.0
Δ13
C
Grainyield(kghaday-1
)
A relation significant only under drought stress)

25. Other traits that get affected by Δ13C
Correlations of carbon discrimination (Δ13
C) with other drought-related traits
observed with the 280 accessions of the reference collection of the chickpea
germplasm in 2008-09.
____________________________________________________________________
Δ13
C
-----------------------------------------------
Characteristics Drought stressed Optimally irrigated
____________________________________________________________________
Days to 50% flowering -0.019 0.103
Days to maturity 0.102 0.147
Shoot biomass (kg ha-1
) 0.153 0.160
Seed yield (kg ha-1
) -0.193*
-0.232*
Harvest index -0.334***
-0.406***
Pod number m-2
-0.383***
-0.273**
Seed number m-2
-0.378***
-0.264**
Seeds pod-1
-0.182 -0.043
100 seed weight (g) 0.321***
0.101
Shoot biomass (kg ha-1
day-1
) 0.091 0.131
Seed yield (kg ha-1
day-1
) -0.263**
-0.207*
___________________________________________________________________

26. The relationship between leaf Δ13C under drought stress with that
under optimally irrigated in the 280 accessions of the chickpea
reference collection in 2008-09.
y = 0.47x - 15.2
r2
= 0.36***
-29.5
-29.0
-28.5
-28.0
-27.5
-27.0
-26.5
-28.5 -28.0 -27.5 -27.0 -26.5 -26.0 -25.5 -25.0
Δ13
C - Drought stressed
Δ13
C-Optimallyirrigated

27. Phenotyping for Harvest index

28. Partitioning Coefficient or Rate of Partitioning
y = 25.1x + 572.0
r2
= 0.72
0
500
1000
1500
2000
2500
0 10 20 30 40 50 60
Harvest index (%)
Grainyieldunderdroughtstress(kgha-1
)
Relation ship of HI with drought yield in CP refernce collection, 2009-10
___________________________________________________________________
Shoot Seed Harvest
Biomass yield index
No Accession (kg ha-1
) (kg ha-1
) (%)
__________________________________
1 ICC 12324 4785 1496 30
2 ICC 8151 4923 1516 29
3 ICC 11879 4754 1517 30
4 ICC 7150 4804 1517 30
5 ICC 5878 3311 1530 52
6 ICC 12866 3237 1538 53
7 ICC 5879 3273 1572 54
8 ICC 13892 3413 1576 51
______________________________________________________________________
E.g. Reference collection
accessions with large HI variation
but not reflecting in yield.
Harvest index in reference collection
is well associated with yield.

29. What happens to the growth duration under ter. drt?
35.1 54.0
35.8 76.5
0 20 40 60 80 100 120
Optimally irrigated
Drought stressed
0 20 40 60 80 100 120
Optimally irrigated
Drought stressed
Vegetative Reproductive
Vegetative Reproductive
ICC 16374
Mean in reference set

30. The value of harvest index under terminal drought
Direct and indirect contributions of HI to Seed yield in the reference
collection of chickpea germplasm in the drought-stressed and
optimally irrigated environments in 2008-09 & 2009-10 seasons
(Association and suggestions of HDU et al. acknowledged)
-0.400
-0.200
0.000
0.200
0.400
0.600
0.800
1.000
1.200
1.400
2008-09-IR 2008-09-NI 2009-10-IR 2009-10-NI
Year x soil water environment
Pathcoefficients
Delta 13C
Days to 50% flr
Days to Maturity
HI
Shoot DM (kg ha-1)
Pod No m-2
100 Seed weight g
Contribution of HI to yield

31. The value of harvest index under terminal drought
Direct and indirect contributions of Pods m-2 to Seed yield in the
reference collection of chickpea germplasm in the drought-stressed
and optimally irrigated environments in 2008-09 & 2009-10 seasons
(Association and suggestions of HDU et al. acknowledged)
-0.400
-0.200
0.000
0.200
0.400
0.600
0.800
1.000
2008-09-IR 2008-09-NI 2009-10-IR 2009-10-NI
Year x soil water environment
Pathcoefficients
Delta 13C
Days to 50% flr
Days to Maturity
HI
Shoot DM (kg ha-1)
Pod No m-2
100 Seed weight g
Contribution of Pod No m-2
to yield

32. Partitioning Coefficient or Rate of Partitioning

33. The relationship of seed yield with partitioning coefficient across 19
(year1)and 40 (year 2) advanced breeding lines grown under drought
stressed and optimally irrigated environments (Krishnamurthy et al. 1999)

34. Yield-based approach
for assessing
Drought Tolerance

35. The drought stressed section of the DT field evaluation
of the reference collection of chickpea germplasm

36. Residuals derived using the multiple regression approach.
This approach considers grain yield under drought stress (Ys)
as a function of yield potential (Yp), time to 50% flowering (F),
and a drought tolerance index (DTI) such that the yield of a
genotype can be expressed as follows:
Ysi = a + bYp + cFi + DTIi + E,
where E is random error with zero mean and variance σ.
Standard residuals (= DTI) were calculated as the difference
between the actual and estimated yields under stress upon
the standard error of the estimated yield (σ).
For this multiple regression, 50% flowering (Fi) under stress for every individual
plot and for the Yield potential (Yp) arithmetic mean across the three replications
were considered. (Bidinger et al., 1987; Saxena et al., 1987; Saxena, 2003; Vadez et al.,
2007; Krishnamurthy et al. 2010; 2011)
Yield-based approach

37. Drought tolerance indices and seed yield at maturity of the highly drought
tolerant and highly drought sensitive mini core chickpea germplasm
accessions.___________________________________________________________________________________________________
S.No Accession Drought Seed Drought Seed Drought Seed
tol. Index yield (kg ha-1) tol. Index yield (kg ha-1) tol. Index yield (kg ha-1)
2002-03 2002-03 2005-06 2005-06 2006-07 2006-07
___________________________________________________________________________________________________
Highly drought tolerant
1 ICC 867 0.71 2472 0.28 924 1.26 1620
2 ICC 1923 0.52 1541 0.57 1059 1.10 1775
3 ICC 9586 0.90 1900 0.31 784 1.18 1328
4 ICC 12947 1.36 2368 0.50 998 0.61 1363
5 ICC 14778 1.11 2570 0.47 1035 1.24 1636
Mean 0.92 2170 0.43 960 1.08 1544
Highly drought sensitive
1 ICC 1052 -0.70 1196 -0.09 642 -0.76 910
2 ICC 2242 -1.10 261 -0.25 226 -1.38 376
3 ICC 2720 -0.86 797 -0.35 213 -1.44 417
4 ICC 2990 -0.62 945 -0.25 431 -0.78 913
5 ICC 3776 -1.35 813 -0.03 719 -0.63 1027
6 ICC 4814 -0.79 943 -0.18 726 -0.64 831
7 ICC 6263 -1.24 1151 -0.04 866 -0.95 1175
8 ICC 6306 -0.41 220 -0.10 229 -1.02 409
9 ICC 7184 -0.87 827 -0.48 457 -1.48 762
10 ICC 7819 -0.42 668 -0.02 458 -1.06 1046
11 ICC 8058 -1.18 973 -0.31 600 -0.82 1057
12 ICC 11764 -0.90 1013 -0.26 429 -0.34 968
13 ICC 11879 -0.93 986 -0.12 702 -0.68 1138
14 ICC 12537 -0.58 1873 -0.23 683 -0.85 1230
15 ICC 12928 -0.82 1140 -0.40 512 -0.68 1130
16 ICC 13124 -1.25 2252 -0.14 1035 -0.55 1753
17 ICC 13441 -0.78 344 -0.02 424 -1.15 598
18 ICC 14669 -0.89 1981 -0.31 788 -0.54 1709
19 ICC 16374 -1.66 1306 -0.42 632 -0.59 1131
20 ICCV 2 -1.26 2102 -0.23 729 -0.60 1403
Mean -0.93 1090 -0.21 575 -0.85 999
___________________________________________________________________________________________________

38. Conclusions
• Many-fold phenotypic variation exists in the reference
collection for roots (structure & function), canopy
temperature, Δ13C, rate of partitioning, drought yield
& DTIs.
• QTLs associated with most drought tolerance traits
(like roots and Δ13C ) appear at a hotspot.
• The rate of partitioning is known to confer yield
advantages under terminal drought; however the GxE
interaction & marker association is yet to be
ascertained

39. Thank
you