Markers-Assisted Selection for Plant Breeding - ILSI India

Markers-Assisted Selection
for
Plant Breeding
P. K. Gupta
Molecular Biology Laboratory
Department of Agricultural Botany
Ch.Charan Singh University, Meerut

MAS vs Transgenics
• MAS should be preferred over transgenic, if
genetic variation is available in gene pool
• MAS is also desirable, if visual selection is
difficult & cost/time ineffective
• Sometimes, both approaches are used
(e.g. insect/disease resistance in rice)

Marker-Assisted Selection for
Conventional Breeding: Why?
- Difficult traits (e.g.,abiotic stresses)
- Traits with low heritability
- Pyramiding of resistance genes
- Selection at seedling stage
- Distinguish homo- & heterozygotes
- No question of biosafety and bioethics

Steps Involved in MAS
• Development of trait-associated markers
• Selection of markers (relative contribut’n)
• Validation of markers (breeding material)
• Utilization of markers (several methods)
• Simultaneous marker develop’t & utilizat’n
(e.g. AB-QTL; ‘mapping-as-you-go’)

Marker Polymorphism for MAS

Marker-Assisted Backcrossing
Transfer of genes from DG to RG
- Foreground selection (for donor’s trait)
- Background selection (for host genome)
(i) 2 step selection (chromosome, genome)
(ii) 3-4 step selection (gene region arm
chromosome genome)

Restricted Backcrossing &
Double Haploidy (DH) for MAS
• BC 1 F 1 pollen for haploids
-gene selection at the haploid stage
-background selection of DHs
• gain over phenotypic selection, and cost
reduced by 40%
(Molecular Breeding 2005)

Some Novel Strategies
(for complex & multiple traits)
• Marker-assisted recurrent selection (MARS)
and genome-wide selection (GWS)
• AB-QTL: transfer of several traits:
Tanksley & Nelson (1996) TAG 92:191
• Introgr’n Lines (ILs) for gene pyramid’g
Zamir (2001) Nat Rev Gen; Ashikari &Matsuoka (2006) Trends Pl Sci
• Breeding by Design TM
Peleman & van der Voort (2003) Trends Plant Sci 8:330

Achievements of MAS
(details not being discussed)
• MAS Programs World-Wide: USA,
Canada, Australia, CIMMYT, IRRI -
Many Cultivars Released
• MAS in India: Cultivars Released
-1. Rice (Improved PB-1; Improved
Sambha Mahsuri; 2. Pearl-millet
(HHB-67-2); 3. Maize (Vivek-QPM9)
• Submergence tolerance: Sub1A

60 Genes/QTL for MAS in Wheat
(Gupta et al. 2009, Mol Breeding)
A
1 2 3 4 5 6 7
QGw.ccsu-1A.1
(1AS-5 0.20)
Glu-A1
(1AL1-0.17-0.61)
Lr46
QGw.ccsu-1A.3
(1AL3-0.61-1.00)
Lr37/Yr17/Sr38
(2AS5-0.78-1.00)
Bx7, Cre 493
PPO (2AL0.77-0.85)
QFhs.ndsu-3AS
(3AS4-0.45-1.00)
QPhs.ccsu-3A.1
(3AL5-0.78-1.00)
PHS, GBSS Null
Yr34 (5AL23-0.87-1.00)
Sr26
GBSS Null
Lr47
(7AS-6 0.21)
Flour Color
Sr22 (7AL 0.83-1.00)
B
Yr10,Yr15,Glu-3
(1BS.sat18-0.50-1.00)
1B/1R
Glu-B1(1BL1-0.47-0.69)
Lr46
(1BL 0.84-0.89)
Lr13/23
Cre1
(2BL 0.50-0.89)
Sr2,Fhb1(3BS-0.78-1)
Rht1(4BS8-0.57-0.81)
Sr36, Sr32,
Sr39/Lr35,
Lr46/Yr29 Cr Yls
Qss.msub-3BL
(3BL-0.81-1.00)
BGGP
GPC-B1,Yr36
(6BS-5 0.49-0.68)
Cre8
Lr9 (6BL-0.79-1.00)
Yfc, Rlnn1, Bo1, LMA
Rht8
(C-2DS 0.47-1.00)
Sr33
Rht2 (4DS3-0.67-0.82)
PinA (5DS7-0.67-0.78)
Lr34/Yr18 (7DS-4 0.61-1.00)
Stb4
D
Glu-D1
(1DL2-0.41-1.00)
Cre3, PPO
(2DL9-0.76-1.00)
Lr24 (3DL-0.78-
Sr24 1) (3DL2-0.27-0.81)
Almt-1
(4DL 0.71–0.86)
Sr30, Lr1
Pch1, Lr19/Sr25,
GBSS Null
Bdv2 (7DS-8 0.77-1.00)
Figure 1

MAS in Cereals
• Maize, Wheat, Barley (CIMMYT,
USA, Australia, Canada)
• Markers for genes for all major traits
(>20 in each case) except yield and
abiotic stresses being used for MAS:
-insect -pest resistance, protein
quality, & other agronomic traits

Gene-Assisted selection (GAS) for
Bread Making Quality
Objective: provide an integrated manipulation of bread
making characteristics using molecular markers
Molecular markers
for gluten strength
STS for Glu-1
New varieties
Molecular markers
for hardness
SNPs for Pinb-D1
Molecular markers for
grain protein content
CAPS for Gpc-B1

MAS for Blight and Blast of Rice
Bacterial Blight
Blast Disease

Pyramiding Xa Genes for Bacterial
Blight Resistance in Rice
• ‘Angke’ & ‘Conde’ (Indonesian cultivars) xa5 in
the background of xa4
Toennissen et al. (2003) Curr Opin Plant Biol 6:191-198
• PR106 (PAU) & MH2R
xa5 (xa5+xa13+Xa21)
(Mahyco Res. Ctr, Jalna)
Singh et al. 2001. TAG (PR106 × IRBB62 xa5 + xa13 + Xa21 ); Mahyco
(MH2R xa5 × IRBB60 Xa4+xa5+xa13+Xa21 )
• PB-1(with basmati quality) with BB resistance
(from IRBB55 xa13+Xa21 ) at IARI
Joseph et al. 2004. Mol Breed 13:377
• Improved Sambha Mahshuri: BLB genes xa5,
xa13, Xa21; donor PR 106; DRR, Hyderabad

Improved Samba Mahsuri (IET 19046)
A first variety from Marker Assisted Selection (MAS)

Bacterial Blight in Rice

Pyramiding of Genes for Blast
Resistance in Rice
• Pi-2(t)+Pi-9(t) in ‘Kalinga III’ &‘Vandana’
(CRRI) * *C101A51 Pi-2(t) & O. minuta Pi-9(t) ;
Pi-1(Pi1)+Pi-2(Piz-5)+Pi-4(Pita) in ‘CO39’
(UAS, Bangalore)
LAC23 Pi-1 , 5173 Pi-2 & Pai-kan-tao Pi-4 → CO39 = CO39 Pi-1+Pi-2+Pi-4
(Hittalmani et al. 2000. TAG 100:1121–1128); *alleles indicated in parenthesis
‘IR50’, ‘Jyothi’(Madras Univ), ‘IR36’, IR50, ‘IR64’ & ‘IR72’
(UAS; Hittalmani 2005)

Resistance for Rice Blast Disease

Submergence Tolerance in Rice
Swarna- Sub1A (Nature, Aug ’06)
BC3F2, BC2F3
Sub1 donor
IR49830
Swarna
six varieties* close to
release at national &
state levels in
Bangladesh & India
*Swarna, IR64,CR1009,
BR11, TDK1, Samba Mahsuri

Marker Assisted Resistance Breeding
Some Often Cited Examples
• Soybean Cyst Nematode
Cregan et al. 2000; Meksem et al., 2001
• Resistance against Gall Midge in Rice
Madan Mohan (ICGEB; Now at UDSC)
Two Hybrids Released in India
• Pearl-millet (HHB-67-2 from HHB-67 (HAU)
(downy mildew resistant): ICRISAT
• Maize (Vivek-QPM9): VPKAS, Almora
Hybrid Rice Cultivars in China
(e.g., Shanyou 63; Restorer Line Minghui 63 Improved)

Gene Pyramiding for Biotic &
Abiotic Stresses in Barley
• South Aust. Barley Impr. Program (SABIP)
- CCN (Ha2/Ha4), BYDV (Yd2), SFNB (Rpt4) &
Mn effi.(Mel1) -‘Sloop’ → ‘Sloop SA’;‘Sloop Vic’
Source: Chebec(Ha2), Franklin (Yd2) & Keel (Ha4, Rpt4 & Mel1)
- CCN, SFNB, leaf rust, leaf scald, boron tol. &
thermostable β-amylase to ‘Gairdner’
Source: 1.‘Keel’– CNN & SFNB resistance, 2. ‘Fanfare’- rust resistance, 3. H. spontaneum
- scab resistance, 4. ‘DH115’ – boron tolerance & 5. SD3 - thermostable β-amylase
SFNB, Spot Form Net Blotch

Problems in MAS for Yield:
Maize/Rice/Tomato/Barley
• Limited success: 20 QTLs in maize † , 2
QTLs in rice ‘9311’ * & 3 QTLs in tomato ** ;
† M. Edwards, Monsanto, St. Louis, USA *O. rufipogon ‘RGC105491’ to O. sativa ‘9311’
(Liang et al. 2004, Euphytica); ** Crossing ILs (S. pennellii → S. lycopersicum ‘M82’),
Gur & Zamir (2004) PLoS Biol 2:e245
• No benefit (epistasis & env.): QTL from
‘Steptoe’ (high yield) & ‘Morex’ (malting) in
Barley → 10 Morex NILs & 1 Steptoe NIL
(Kandemir et al., 2000 Mol Breed 6:157)

Pyramiding Additional Genes
Over Transgenes
• Rice: ‘CO39’NIL Piz-5 →IR50 Piz-5 → transform Xa21 =
IR50 Piz-5+Xa21 (Crop Sci 42:p2072, 2002)
• Rice: IRBB21 Xa21 → Minghui 63 Xa21 (MAS) × Minghui
63 cry1Ab+cry1Ac (GM) = Minghui 63
cry1Ab+ cry1Ac+Xa21
(Minghui 63 is a widely used cms restorer line for hybrid rice in
China)
(Jiang et al. 2004 Plant Breed 123:112-116)
• Soybean: QTL for insect resistance from ‘PI
229358’ to GM ‘Jack-Bt’ cry1Ac
(Walker et al. 2002; Mol Breed 9:43)

Reasons for Low Impact &
Future Needs for MAS
• Non-availability of robust markers
• Epistasis, background and g × e
• Still cost-ineffective: high throughput & low
cost genoytpic centres needed
• Plant breeder-molecular biologist gap
• Association mapping needed
• MARS & GWS for complex/multiple traits

Further Reading on MAS
• Gupta et al. (2009). MAS for conventional
plant breeding. Plant Breed Rev 33:145-217
• Gupta et al. (2009). MAS in Wheat: Mol
Breeding (Special Issue): 24: (published online,
11 December, 2009)
• Gupta et al. (2009). MAS for complex and
multiple traits in crops. (under preparation)
• Several Other Reviews on this Active Area