Application of Molecular Tools for Wheat Quality Improvement: Indian Scenario

H. S. Balyan Ch. Charan Singh University, Meerut

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

Wheat Grain Quality

1. 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 Quality

4. 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
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

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)

PAU & IARI

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

Framework Molecular Maps

GPC Map: 214 loci (CCSU &NCL) GW Map:298 loci (CCSU) PHST Map:214 loci (CCSU) GH Map: Under preparation (IARI)
M WL711 PH132 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15

Prasad et al. (2003). TAG 106:659-667

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

QTL Analysis for GPC

Composite Interval Mapping (CIM) (QTL Cartographer)

- 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
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

CSSU

Validation of SSR markers Linked With QTLs for GPC in NILs (CCSU)
Xgwm830 Xgwm359 Xgwm1198 Xgwm1115 Xgwm1052 Xgwm515 Xgwm372 Xgwm895 Xgwm164 Xgwm357 Xgwm456 Xgwm892 Xgwm374 Xgwm429 Xgwm410 Xgwm319 Xgwm1249 Xwmc272 Xgwm601 Xgwm133 Xgwm771 Xgwm680 Xgwm889 Xgwm1171 Xgdm86

C
Xwmc41 Xgwm608 Xgwm1264 Xgwm1204

C
Xgwm1063

C
Xgwm894 Xgwm397

C
Xgwm940b Xgwm513

C
Xgwm831

C
Xgwm805

Xgwm957

Xgwm1256

Xgwm1027

1A

1D

2A

2B

2D

3A

3D

4A

4B

5B

5D

6B

7A

7D

NIL2233

Xgwm493

Xgwm493

Xgwm726

Xgwm493

Xgwm830

Xgwm456

C
Xgwm374 Xgwm429 Xgwm410 Xgwm319 Xgwm1249 Xwmc272

C
Xgwm940b Xgwm513

Xgwm429 Xgwm410

C
Xgwm429 Xgwm1249

C
Xgwm940b

Xgwm1027

Xgwm666

Xgwm1027

2B

3B

3D

4B

2B

3B
NIL2215

5A

2A

2B

3B

4B

2A
NIL2232

NIL2210

NIL2218

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
Kumar et al. (2006). Euphytica 151:135-144
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

CSSU

QTL Analysis for PHST CIM

7 QTLs on 1A, 2A, 2D, 3A, 3B One major QTL each on 2A and 3A

HD2329

SPR8198

Kulwal et al. (2005). TAG 111: 1052-1059

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

CSSU

Two-Locus QTL Analysis QTLMapper

PVE due to M-QTLs: GPC=7.22 to 7.22%; GW=28.11%; PHST=37 to 48% PVE due to QQ: GPC= 2.68 to 6.04%; GW=42.12%; PHST=27 to 28.73% PVE due to QE: 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

Q Q and Q E Interactions Resolved Through Two-Locus QTL Analysis for GPC in Bread Wheat
1A 1B 1D 2A 2B 2D 3A 3B 3D 4A 4B 4D 5A 5B 5D 6A 6B 6D 7A 7B 7D

:a

: ae

: a + ae

: aa

: aae

: aa + aae

: centromere

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

CSSU

Linked Markers and Their Validation

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, E35M4718 Leaf rust: Lr9 (SCS5550); Lr24(SCS73719); Lr28(SCS421570); Lr19 (SCS265512); (SCS253736)
CSSU

Strategy for MAB

Three step selection during backcrossing - Foreground selection for alleles for donorparent markers flanking the QTL - Background selection for the recipientparent-alleles on loci of individual chromosome carrying the QTL - Background selection for the recipientparent-alleles for the whole-genome
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

CSSU

MAB of GPC-B1,a high GPC QTL, into 10 Indian Wheat Genotypes

10 Recipient Parents (RP)
Indian cultivars

High GPC Donor Parent (DP)

Yecora Rojo

Fs

Respective RP

BC1F1S Foreground & background selections BC3F1s BC3F2s Selection of individuals homozygous for GPC-B1a DP RP BC3F2 progenies (85-100% genomic similarity)

Xuhw89

**

Asterisk indicate individuals homozygous for the GPC-B1 allele

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

*
CSSU

Protein content (%)

HD2 6 87

HD2 329
PBW 343

PBW 373

PBW3 43(Lr1 9)
PBW34 3(Lr9)

(Change in GPC by -0.93 to 1.72%)

PBW3 43(Lr2 4)

HI977
Raj376 5

Performance of BC3F1 Containing GPC-B1 QTL Allele

K910 7

10
CSSU

11

12

13

Summary of MAS for Gpc-B1

Generati on F1 BC1F1 BC2F1 BC2F2 F4 BC2F3/F5 Year 2004 2004-05 2005-06 2006-07 2006-07 2007 Total plants analysed 167 330 300 ~50 DNA isolation +ve for ASA 74 110 168 25 In progress Location Keylong PAU, Ludhiana Keylong PAU, Ludhiana PAU, Ludhiana Keylong

Recipient Parents- BW3652, C273, C518, DBW16, DBW17, BW4406, BW4426, PBW509, PBW554, PBW563, PBW564, PBW568, PBW571

PAU

Grain Protein content of F4s progenies of the crosses of GluPro with wheat
20
15.6% 13.0 to 14.3%

15
11.3%

%GPC

10
PBW343

5 0

Marker assisted selection for HGPC gene GpcB1 using allele specific amplification marker (ASA)

Glupro

PAU

PHS scale

PHS scale Susceptible =9 Resistance =1

1 BC3F

HD2 329 (RP )

SPR

8198 (DP)

Parents and Progeny

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

PBW343 X F1 Half seed analysis BC1F1 Half seed analysis BC2F1 Half seed analysis BC2F2 Half seed analysis BC2F3(2006-07) X PBW343 Half seed analysis BC3F1 BC3F2(2006-07) X PBW343 X T. durum-T. dicoccoides PBW343

~300 half seeds analysed and 8 homozygous and 11 segregating progenies identified

PAU

Iron and zinc of wild relatives as compared with cultivars

100 90 80 70 60 50 40 30 20 10 0
70 91 07 0 3 4 2 2 3 34 PB W T. du ru m
IARI, ARI, PAU, IITR

Iron and Zinc (m g/kg)

64

57

77

77

37

37

35

77

sc hy

sc hy

de

ss im

eg

ss im

ri n

ri n

co i

co i

de

.k ot

.k ot

eg

gi

gi

oc

.l on

.l on

.p

er

oc

Ae

Ae

.p

dic

dic

Ae

Ae

Ae

T.

Ae

T.

Iron (mg/kg)

Zinc (mg/kg)

Accessions

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

T.

ae

st

iv u

er

PD W

s4

s4

i3

i3

13

27

Spikes of Parents and F1 of wide crosses

Chinese Spring

(PhI)

F1 CS (PhI) X Ae. kotschyi

Ae.kotschyi
IARI, ARI, PAU, IITR

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

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

e
ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

f
IARI, ARI, PAU, IITR

Synthetic amphiploids

AABBDD

ABDUS

UUSS

Chinese Spring (PhI)

F1 CS (PhI) X Ae. kotschyi

Ae.kotschyi

Sterile

Colchicine treatment

AABBDDUUSS

Synthetic amphiploid

IARI, ARI, PAU, IITR

Grain Iron and Zinc of Amphiploids

Parents/ Amphiploid WL 711 CS (Phi) Ae. kotschyi 3774 Ae. kotschyi 3790 Ae. kotschyi 396 WL711 x Kot 3774 CS(Phi) x Kot 396 WL711 x Kot 391 CS(Phi) x Kot 3790 Fe (ppm) 30.87 30.84 82.67 74.55 77.00 71.96 70.73 70.00 77.27 Zn (ppm) 17.23 19.61 41.22 41.67 51.07 25.20 33.68 33.13 32.75

ICAR-ACIAR Workshop on Molecular Marker Assisted Breeding In Wheat, October 12, 2007

IARI, ARI, PAU, IITR

Leaf iron and zinc content in sterile F1 hybrids between wheat and Aegilops donors
180 160 140 Ir nc n n (m /k ) o o te t g g 120 100 80 60 40 20 0 a b c d e f g h i j k l m n o
Donor

Re cipie nts, F1 hybrids, Donors

Recepient

F1 Hybrid

70 60 Z cc n n (m /k ) in o te t g g 50 40 30 20 10 0

m
F1

o
Donor

Re cipie nts , F1 Hybrids , Donors Recipient

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