Molecular breeding & genetic engineering
for drought tolerance
What scope ?
Vincent VadezICRISAT
ICRISAT in shortInternational Crops Research Institute for the Semi-Arid Tropics
5 mandate crops: Chickpea, pigeonpea, groundnutPearl millet, sorghum
1.3 Billions people in drought-prone areas (India/Africa)Drought is a major research focus for ICRISAT
Drought research at ICRISAT
Today’s presentation
MAS (pearl millet, sorghum)- PNHI and staygreen
QTL identification (chickpea)-Roots
QTL identification + Transgenics (groundnut)- Transpiration Efficiency (TE)
MAS for pearl millet
Pearl millet line-source irrigation experiment, summer drought nursery, ICRISAT-Patancheru
MAS for pearl millet
LG 2M 322
M 394
M 214
M 321M 592 M 443M 356
M 738
Panicle no.2
Biomass yield2
Stover yield2
Drought tolerance
per se
Stress performance
Stressperformance
“H 77 alle le” “PRLT allele”
40 cM
Grain yield2
100-grain mass3
Harvest index3
Biomass yield1
Grain yield3
100-grain mass3
Harvest index3
Panicle grain no.1
Leaf rolling1
A major terminaldrought tolerance QTL on LG2
Gra
in y
ield
(g m
-2)
250
270
290
310
330
350
370
390
410
430
450
100% 92% 83% 75% 66% 58% 49% 41% 32%
Percentage of non-stress irrigation
ICMR 01029PRLT 2/89-33H 77/833-2
MAS for pearl millet
Near-isogenic pearl millet hybrids differing in terminal drought tolerance
843A x ICMR 01029
843A x H77 (=HHB 67): Smaller percentage of florets setting grains: Poorer grain filling
MAS for pearl millet
Neutron probe observations in the field
20
25
30
35
Moi
stur
e at
90-
105
cm
PRLT2/89-3H77/833-2
ICMR 01029
Dates of sampling
Grain filling
Tolerant material deplete more moisture at deeper layers
MAS for sorghum
0
20
40
60
80
100
120
1 2 3 4 5 6 7 8 9
Weeks from flowering
% G
reen
leaf
are
a
ISIAP DoradoIDSG 02034IDSG 02368
Staygreen Senescent
Maintenance of a green leaf areaUnder terminal drought
MAS for sorghum
Evaluation in the field under terminal drought
0.600
0.650
0.700
0.750
0.800
0.850
0.900
0.950
1.000
0.000 0.100 0.200 0.300 0.400 0.500 0.600 0.700 0.800
Rel
ativ
e 10
0 G
rain
Mas
s
R2 = 0.72
Relative %GLA
Roots – staygreen contribute to better grain filling
MAS for sorghum
ISIAP Dorado
Range in IL ILs >Dorado
ILs <Dorado
Harvest index (%) 40.6 28.3 − 44.4 2 11Grain yield (g plant-1) 54.2 34.5 − 57.4 0 13Stover yield (g plant-1) 65.8 41.4 − 77.0 1 5
NIRS-predicted NDM and digestibility values of stoverEntry name NDM, % of
R 16 meanDigestibility,
% of R 16 mean
RSG 03061 147% 108%
RSG 03123 137% 118%
Need to advance more generations of backcrosses
MAS for sorghum
6 QTLs related to the stay green character Staygreen consensus QTL of B35
LgB LgC LgD LgG LgH LgILgA LgE LgF LgJ
Linkage drag (dwarfing genesGrain setting and quality)Stg2
Stg1
StgB
Stg3
StgA
Stg4
Stg2LgC
Xtxp231
Xtxp59
Xtxp31
Xtxp120
R locus (pericarppigmentation)
Z locus (mesocarpthickness)
Need to further saturate the staygreen QTL genome portion
MAS for sorghum
Evaluation in large 2.0m PVC cylinders under drought
05
1015202530
30-60
cm90
-120 c
m15
0-180
cm21
0-240
cm27
0-300
cm
% o
f roo
t in
shal
low
laye
r
R16DoradoB35E36-1
Staygreen
Roots are involved in staygreen QTL(s)
Chickpea - RootsBefore
Chickpea - Roots
Screening of the minicore collection for root depth
Dep
th (c
m)
140
020406080
100120
160
Screening procedure easier (1.2 m PVC tubes)
Chickpea - Roots
r = 0.77
RLD at 60cm depth
Seed
yie
ld
A good relation between deeper rooting and seed yield
An essential question to answer: Roots for what environment??
Chickpea - RootsICC283
0.70
0.80
0.90
1.00
1.10
234 Genotypes tested
Roo
t Dep
th (m
)
Annigeri ICC4958 ICC8261
One QTL from “old” population Annigeri x ICC4958
New RIL pops from more varied parents
About 350 SSR markers available: Need more!!
Groundnut - Transpiration efficiency
Two approaches:
RILs DREB1A transgenics
Groundnut - Transpiration efficiency
Phenotyping of RIL for transpiration efficiency (TE) in groundnut
RILs
Groundnut - Transpiration efficiency
RILs
0
1020
3040
50
2.65 2.75 2.85 2.95 3.05 3.15 3.25 3.35 3.45 3.55
Transpiration efficiency (g kg-1)
Freq
uenc
y of
RIL
s (n
umbe
r)
TAG 24
ICGV86031
Good segregation between RILs
Groundnut - Transpiration efficiency
Available SSR markers : 900
Already tested : 463
Polymorphic markers : 35
18
E7
2C
11
20
12
13
E0
9
T I T I T I T I
Segregation data on ABI (9 markers)
T= TAG24, I= ICGV 86031AA
A
B
B
B
A clear need todevelop more markers
Develop syntheticgroundnut from wildancestors
RILs
Groundnut - Transpiration efficiency
Testing TransgenicsTesting Transgenics
P2 Glasshouses
DREB1A
Groundnut - Transpiration efficiency
DREB constructs used for DREB constructs used for AgrobacteriumAgrobacterium--mediated transformation
DREB1Amediated transformation
Groundnut Variety Used: JL 24• Total no. of transgenic events: 30• Generation: T3 to T5
Groundnut - Transpiration efficiencyGroundnut Dry-Down set-up
DREB1A
Groundnut - Transpiration efficiency
Wild type JL24DREB1A
Dendrogram based on soil moisture value where Trdeclines and number of days to deplete soil water
RD19
RD12
RD20
RD2RD11
Groundnut - Transpiration efficiency
TE under well watered and drought stressin 5 DREB1A events and JL24
DREB1A
JL 24RD 19RD 12RD 20RD 2RD 11
2.054.315.133.194.094.96
TE - WW TE - DS
4.294.994.634.526.125.59
Most DREB1A have larger TE than JL24 under WWTwo events with higher TE under both water regimes
Groundnut - Transpiration efficiency
Range of values in TE in a RIL population and in DREB1A groundnut
DREB1A
01234567
RIL DREB WellWatered
DREB drought
TE (g
DW
/kg
wat
er)
Min
Max
Larger range of variation for TE in DREB1A
Conclusion
MAS (pearl millet and sorghum)Need to saturate QTL areas to increase the efficiencyPyramid with other QTLsGet rid of linkge drag
Root-Chickpea: More markers, QTL identification, Start marker-assisted breeding
TE-Groundnut: More markersRe-synthesize the cultigen from its ancestorsTransgenic approachNew traits
Thank you