application of molecular tools for wheat quality ...aciar.gov.au/files/node/3875/session...
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Application of Molecular Tools for Wheat Quality Improvement: Indian Scenario
H. S. BalyanCh. Charan Singh University,
Meerut
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
Wheat Grain Quality1. GPC & micronutrients (Fe, Zn)
- Nutritional quality
2. Pre-harvest Sprouting (PHS)- Activates hydrolyzing enzymes- Endosperm constituents are broken down- Decreased volume & poor crumb of bread- Lowers quality of grain for bread/feed and seed
3. GW- Aesthetic, market & industrial quality (flour recovery)
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
Wheat Grain Quality4. GPC, GW, sedimentation value, kernel
hardness, amylose content- Bread, biscuit, chapati and noodle quality
5. HMW-GS and LMW-GS- Flour processing properties/dough quality such as viscoelasticity & extensibility
6. Phytic acid- Reduces absorption of Fe and Zn
Applications of Molecular Markers
- Preparation of genetic maps- QTL analysis- High density mapping- Validation of linked markers- Molecular marker assisted selection
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
RIL Populations Used
- RIL pop for GPC (HD2329 × PH132)- RIL pop for PHST (WL711× SPR8198)- RIL pop for GW (CS × Rye Selection111)- RIL population for GH (K65 × Lerma Rojo)
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
PAU & IARI
Framework Molecular Maps GPC Map: 214 loci (CCSU &NCL)GW Map:298 loci (CCSU)PHST Map:214 loci (CCSU)GH Map: Under preparation (IARI)
Prasad et al. (2003). TAG 106:659-667
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
M 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15PH13
2
WL7
11
- 10 QTLs for GPC detected (LOD 2.5/above threshold LOD)
- 9 chromosomes: 2A, 2B, 2D, 3D, 4A, 6B & 7A
- Range of PVE: 2.95% to 32.44%Prasad et al. (2003). TAG 106: 659-667
Composite Interval Mapping (CIM) (QTL Cartographer)
QTL Analysis for GPC
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 CSSU
Xgwm830
Xgwm1063Xgwm164Xgwm357
2A
Xgwm359Xgwm1198
Xgwm1256
Xgwm1052Xgwm515Xgwm372
Xgwm895
Xgwm1115
C
1D
C
Xgwm957
Xgwm608
2B
Xgwm319Xgwm1249
Xgwm374
Xwmc272
Xgwm1027
Xgwm429Xgwm410
C
2D
CXwmc41
Xgwm1264Xgwm1204
C
3A
Xgwm1171
C
7A6B
Xgwm680
Xgwm133Xgwm771Xgwm889C
Xgwm456
3D
Xgwm892C
Xgwm894
Xgwm601
Xgwm397
C
4A
Xgwm940b
4B
C
Xgwm513Xgwm805
C
5D5B
Xgwm831
C
7D
C
NIL2233
Xgdm86
C
Xgwm666
2B
CXgwm429Xgwm410
Xgwm1027
3B
C
Xgwm493
5A
C
NIL2210
Xgwm456
Xgwm493
2B
CXgwm429Xgwm410Xgwm319
3D
C
Xgwm1249
Xgwm374
Xwmc272
3B 4B
C
Xgwm513
C
2A
C
Xgwm830
NIL2215
Xgwm940b
2A
Xgwm940b
NIL2218
Xgwm1027
2B
Xgwm429C
Xgwm1249
3B
C
Xgwm493Xgwm726
C
4B
C
NIL2232
1A
Validation of SSR markers Linked With QTLs for GPC in NILs (CCSU)
ICIM mapping of QTLs for GW using IciMapping v1.4
• 34 QTLs on 18 chromosomes
• 3 major QTLs detected
• QTL on1B explained 26.0 to 82.0% and another QTL explained 10.6- 45.7 % PV
• QTL on 2B explained 11.37-32.77 % PV
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
Kumar et al. (2006). Euphytica 151:135-144CSSU
QTL Analysis for PHSTCIM
• 7 QTLs on 1A, 2A, 2D, 3A, 3B
• One major QTL each on 2A and 3A
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 CSSU
HD2329 SPR8198 1 2 3 4
5 6 7 8 9
Kulwal et al. (2005). TAG 111: 1052-1059
Two-Locus QTL AnalysisQTLMapper
PVE due to M-QTLs:GPC=7.22 to 7.22%; GW=28.11%; PHST=37 to 48%
PVE due to Q×Q:GPC= 2.68 to 6.04%; GW=42.12%; PHST=27 to 28.73%
PVE due to Q×E:GPC= 25.00 to 47.99% ; GW= nil; PHST= 3.24%
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 CSSU
1A1B1D2A2B2D3A3B
3D4A4B4D5A5B5D6A6B6D7A7B7D
: a : a + ae: ae : aa : aae : aa + aae : centromere
Q × Q and Q × E Interactions Resolved Through Two-Locus QTL Analysis for GPC in Bread Wheat
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 CSSU
GPC: gwm1264, gwm608, gwm1171, gwm508, gwm193, gwm361, gwm644, PCR-CAPS marker, ASA marker, uhw89
PHST: cdo795, psr115, bcd1431.1, bcd1431.2, bcd1380, fbb370, bcd907, fbb283, ATPase.2,wmc104, Mst101,wmc153,gwm155,GW: wmc333, gwm419,E35M4714, E35M4718Leaf rust: Lr9 (SCS5550); Lr24(SCS73719); Lr28(SCS421570); Lr19 (SCS265512); (SCS253736)
Linked Markers and Their Validation
CSSU
Strategy for MAB• Three step selection during backcrossing
- Foreground selection for alleles for donor-parent markers flanking the QTL
- Background selection for the recipient-parent-alleles on loci of individual chromosome carrying the QTL
- Background selection for the recipient-parent-alleles for the whole-genome
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007CSSU
MAB of GPC-B1,a high GPC QTL, into 10 Indian Wheat Genotypes
Yecora Rojo
F1 s ×
Foreground & background selections
BC3F1s (85-100% genomic similarity)
BC3F2s Selection of individuals homozygous for GPC-B1a
Indian cultivars
Respective RP
BC1F1S
10 Recipient Parents (RP) High GPC Donor Parent (DP)×
×
Xuhw89
DP RP BC3F2 progenies
* ** * * * *ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
Asterisk indicate individuals homozygous for the GPC-B1 allele
CSSU
10
11
12
13Pr
otei
n co
nten
t (%
)
HD
2687
HD
2329
PBW
343
PBW
343(
Lr19
)
PBW
343(
Lr9)
PBW
343(
Lr24
)
K91
07
PBW
373
Raj
3765
HI9
77
Performance of BC3F1 Containing GPC-B1 QTL Allele(Change in GPC by -0.93 to 1.72%)
CSSU
Summary of MAS for Gpc-B1
Keylong--2004F1
2007
2006-07
2006-07
2005-06
2004-05
Year
PAU, Ludhiana25~50F4
PAU, Ludhiana74167BC1F1
PAU, Ludhiana168300BC2F2
KeylongIn progress
DNA isolation
BC2F3/F5
Keylong110330BC2F1
Location+ve for ASA
Total plants analysed
Generation
Recipient Parents- BW3652, C273, C518, DBW16, DBW17, BW4406, BW4426, PBW509, PBW554, PBW563, PBW564, PBW568, PBW571
PAU
20
15
10
5
0
Grain Protein content of F4s progenies of the crosses ofGluPro with wheat
%GP
C
PBW
343
Glup
ro
13.0 to 14.3%
11.3%
15.6%
Marker assisted selection for HGPC gene GpcB1 using allele specific amplification marker (ASA)
PAU
PHS
scal
e
Parents and Progeny0
3
6
9
HD23
29 (R
P)
BC3F
1
SPR8
198
(DP)
PHS scaleSusceptible =9 Resistance =1
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 CSSU
Graphical genotypes of PBW343/SPR5 introgression lines
Xwmc153 and Xgwm155 suitable for marker assisted mobilization of PHST to elite wheat backgrounds
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 PAU
Transfer of new GluAx and GluAy subunits to bread wheat
X
BC2F3(2006-07)
Half seed analysis
X
X
X
Half seed analysis
Half seed analysisHalf seed analysis
Half seed analysis
T. durum-T. dicoccoidesPBW343
F1 PBW343
BC1F1 PBW343
BC2F1PBW343
BC3F1BC2F2
BC3F2(2006-07)
~300 half seeds analysed and 8 homozygous and 11 segregating progenies identified PAU
0102030405060708090
100
Ae. ko
tschy
i 357
3Ae.
kotsc
hyi 3
774
Ae. pe
regrin
a 379
1
Ae. pe
regrin
a 137
72
Ae. lon
gissim
a 3770
Ae. lon
gissim
a 3507
T. dico
ccoide
s 464
0
T. dico
ccoide
s 477
2
T. aes
tivum PBW 34
3
T. duru
m PDW 274
Accessions
Iron
and
Zinc
(mg/
kg)
Iron (mg/kg) Zinc (mg/kg)
Iron and zinc of wild relatives as compared with cultivars
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 IARI, ARI, PAU, IITR
Chinese Spring (PhI) F1 CS (PhI) XAe. kotschyi
Ae.kotschyi
Spikes of Parents and F1 of wide crosses
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 IARI, ARI, PAU, IITR
a b
dc
fe
Meiotic chromosome pairing in F1 of wide crosses
CS (PhI) X Aegilops kotschyi 3573
(a) 2II (b) 4II
CS (PhI) X Aegilops kotschyi 396
(c) 4II (d) 6II
CS (PhI) X Aegilops kotschyi 3790),
(e) 3II;,1III (f)2II
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 IARI, ARI, PAU, IITR
Sterile
Colchicine treatment
Synthetic amphiploid
AABBDDUUSS
Chinese Spring (PhI) Ae.kotschyiF1 CS (PhI) XAe. kotschyi
UUSSABDUSAABBDDSynthetic amphiploids
IARI, ARI, PAU, IITR
Grain Iron and Zinc of Amphiploids
41.6774.55Ae. kotschyi 3790
17.2330.87WL 71119.6130.84CS (Phi)
32.7577.27CS(Phi) x Kot 379033.1370.00WL711 x Kot 39133.6870.73CS(Phi) x Kot 39625.2071.96WL711 x Kot 377451.0777.00Ae. kotschyi 396
41.2282.67Ae. kotschyi 3774
Parents/ Amphiploid Fe (ppm) Zn (ppm)
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 IARI, ARI, PAU, IITR
0
20
40
60
80
100
120
140
160
180
a b c d e f g h i j k l m n o
Recipients, F1 hybrids, Donors
Iron
cont
ent (
mg/
kg)
Recepient F1 Hybrid Donor
0
10
20
30
40
50
60
70
a b c d e f g h i j k l m n o
Recipients, F1 Hybrids, Donors
Zinc
con
tent
(mg/
kg)
Recipient F1 Donor
Leaf iron and zinc content in sterile F1 hybrids between wheat and Aegilops donors
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007 IARI, ARI, PAU, IITR
Future Research Programmes to Identify Novel QTLs & Linked Markers
• AB-QTL analysis• LD and association analysis• Identification of novel positive alleles• Transciptome analysis • Expression QTL (e-QTL) analysis• Genetic mapping of newly identified gene
sequences including regulatory sequences
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007
Participating Institutions
• CCSU, Meerut• IARI, New Delhi• PAU, Ludhiana• DWR, Karnal• ARI, Pune• NCL Pune• IIT, Roorkee
ACKNOWLEDGEMENTS
Funds provided by DBT and NATP are gratefully acknowledged
Thank You