Molecular breeding & genetic engineering for drought tolerance ...

mustardnimbleΒιοτεχνολογία

11 Δεκ 2012 (πριν από 4 χρόνια και 8 μήνες)

158 εμφανίσεις

Molecular breeding & genetic engineering
for drought tolerance
What scope ?
Vincent
Vadez
ICRISAT
ICRISAT in short
International Crops Research Institute for the Semi-Arid Tropics
5 mandate crops: Chickpea, pigeonpea, groundnut
Pearl 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

2
M32
2
M39
4
M214
M32
1
M59
2

M44
3
M356
M738
Pani
cle no.2
Bioma
ss yield2
Stov
er yield2
Dr
o
u
ght

tol
er
an
ce
per se
Stress
performance
St
res
s
performance

H
77
a
lle
le”
“PR
LT
all
ele”
40 cM
Grain yi
el
d2
100-grai
n
mass3
Harv
est
i
n
dex3
Bi
o
mass
yield1
Grain yi
el
d3
100-grai
n m
ass3
Harvest index3
Panicle
grain no.1
Leaf rol
l
ing1
A major terminal
drought tolerance
QTL on LG2
Grain yield (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 01029
PRLT 2/89-33
H 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
Moisture at 90-105 cm
PRLT2/89-3
H77/833-2
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
12
345
67
89
Weeks from flowering
% Green leaf area
ISIAP Dorado
IDSG 02034
IDSG 02368
Staygreen
Senescent
Maintenance of a green leaf area
Under 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
Relative 100 Grain Mass
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
11
Grain yield (g plant-1)
54.2
34.5 −
57.4
0
13
Stover yield (g plant-1)
65.8
41.4 −
77.0
1
5
NIRS-predicted NDM and digestibility values of stover
Entry name
NDM, % of
R 16 mean
Digestibility,

% 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
LgI
LgA
LgE
LgF
LgJ
Linkage drag
(dwarfing genes
Grain setting and quality)
Stg2
Stg1
St
gB
Stg3
StgA
Stg4
Stg2
LgC
Xtxp231
Xtxp59
Xtxp31
Xtxp120
R
locus (pericarp
pigmentation)
Z
locus (mesocarp
thickness)
Need
to
further
saturate
the
staygreen QTL genome
portion
MAS for sorghum
Evaluation in large 2.0m PVC cylinders under drought
0
5
10
15
20
25
30
30-60 cm
90-120 cm
150-180 cm
210-240 cm
270-300 cm
% of root in shallow layer
R16
Dorado
B35
E36-1
Staygreen
Roots are involved in staygreen QTL(s)
Chickpea -
Roots
Before
Chickpea -
Roots
Screening of the minicore collection for root depth
Depth (cm)
140
0
20
40
60
80
100
120
160
Screening procedure easier (1.2 m PVC tubes)
Chickpea -
Roots
r = 0.77
RLD at 60cm depth
Seed yield
A good relation between deeper rooting and seed yield
An essential question to answer: Roots for what environment??
Chickpea -
Roots
ICC283
0.70
0.80
0.90
1.00
1.10
234 G
e
no
ty
p
e
s te
ste
d
Root Depth (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
10
20
30
40
50
2.
65
2.
75
2.
85
2.
95
3.
05
3.
15
3.
25
3.
35
3.
45
3.
55
T
rans
p
i
rat
i
o
n e
ffi
c
i
e
nc
y (
g
k
g
-1)
Frequency of RILs
(number)
TAG
24
IC
G
V
86031
Good
segregation
between
RILs
Groundnut -
Transpiration efficiency
Available SSR markers : 900
Already tested
: 463
Polymorphic markers : 35
18E7
2C11
2012
13E09
T I
T I T I T I
Segregation data on ABI (9 markers)
T= TAG24, I= ICGV 86031
A
A
A
B
B
B
A clear
need
to
develop
more markers
Develop
synthetic
groundnut
from
wild
ancestors
RILs
Groundnut -
Transpiration efficiency
Testing Transgenics
Testing Transgenics
P2 Glasshouses
DREB1A
Groundnut -
Transpiration efficiency
DREB constructs used for
DREB constructs used for
Agrobacterium
Agrobacterium
-
-
mediated transformation
DREB1A
mediated transformation
Groundnut Variety Used: JL 24

Total no. of transgenic events: 30

Generation: T3 to T5
Groundnut -
Transpiration efficiency
Groundnut Dry-Down set-up
DREB1A
Groundnut -
Transpiration efficiency
Wild type JL24
DREB1A
Dendrogram
based on soil moisture value where Tr
declines and number of days to deplete soil water
RD19
RD12
RD20
RD2
RD11
Groundnut -
Transpiration efficiency
TE under well watered and drought stress
in 5 DREB1A events and JL24
DREB1A
JL 24
RD 19
RD 12
RD 20
RD 2
RD 11
2.05
4.31
5.13
3.19
4.09
4.96
TE -
WW
TE -
DS
4.29
4.99
4.63
4.52
6.12
5.59
Most DREB1A have larger TE than JL24 under WW
Two 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
0
1
2
3
4
5
6
7
RI
L
DRE
B W
ell
Wa
te
r
e
d
DREB dr
ought
TE (g DW/kg water)
Mi
n
Ma
x
Larger range of variation for TE in DREB1A
Conclusion
MAS (pearl
millet
and
sorghum)
Need
to
saturate
QTL areas
to
increase
the
efficiency
Pyramid
with
other
QTLs
Get
rid
of
linkge
drag
Root-Chickpea:
More markers,
QTL identification,
Start
marker-assisted
breeding
TE-Groundnut:
More markers
Re-synthesize
the
cultigen
from
its
ancestors
Transgenic
approach
New
traits
Thank you