GCP 2011General Research

Meeting

21-25 Sep 2011

L Krishnamurthy

ICRISAT

Chickpeas - Phenotyping for drought traits

• 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

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

• 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

•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

Variation in rooting depth R

oo

tin

g d

epth

(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

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

0.32R

oo

t le

ng

th d

ensi

ty (

cm c

m-3

)

ICC8261

ICC4958

Annigeri

ICC283

ICC1882

SEd=0.022

ICC

M0

24

9

TA

A1

70

GA

24

S

TM

S11

CID traits

Root traits

ICC

M0

24

9

TA

A1

70

GA

24

S

TM

S11

Root + CID + Yield traits

Hot spot for QTL –LG 5 (ICC 4958 × ICC 1882 map)

Root function – Transpiration measurements

Dynamic obs.

Root function

i.e. actual transpiration

Root phenotyping

Are there simple solutions?

Root traits – How useful is the seedling expression?

Contrasting selections of 10 day-old chickpea seedlings from the

reference collection - sand culture.

- for a high throughput assessment of canopy temperature or root function

L. Krishnamurthy, Z.A. Mainassara, J. Kashiwagi, R. Purushothaman &

V.Vadez

Infrared thermography

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

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.

Variation in canopy temperature across 60 drought tolerant

and 24 sensitive genotypes at 75 DAS, 2009-10.

272829303132333435

1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76

Germplasm accessions

C.

are

a x

C.

tem

p.

(0C

)

25

27

29

31

33

35

37

39

1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76

Ca

no

py

te

mp

. (0

C)

0.8

0.9

1

1 6 11 16 21 26 31 36 41 46 51 56 61 66 71 76

Can

op

y p

rop

ort

ion

Evap. 5.1mm

Max T. 30.0 0C

Min T. 9.0 0C

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

Phenotyping for TE (through Δ13

C)

How does TE contribute

to drought tolerance?

Yield = Shoot biomass x Harvest index

Transpiration Transpiration Efficiency (TE)=13C x

Leaf biomass + Stem biomass

LA x Leaf 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

… and the optimally irrigated section of the DT field evaluation

of the reference collection of chickpea germplasm

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

___________________________________________________________________

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

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

Gra

in y

ield

(k

g h

a d

ay

-1)

A relation significant only under drought stress)

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*

___________________________________________________________________

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

Δ1

3C

- O

pti

mall

y i

rrig

ate

d

Phenotyping for Harvest index

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 (%)

Gra

in y

ield

un

de

r d

rou

gh

t s

tres

s (

kg

ha

-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.

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

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

Path

co

eff

icie

nts

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

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

Path

co

eff

icie

nts

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

Partitioning Coefficient or Rate of Partitioning

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)

Yield-based approach

for assessing

Drought Tolerance

The drought stressed section of the DT field evaluation

of the reference collection of chickpea germplasm

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

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

___________________________________________________________________________________________________

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

Thank

you