1

APPLICATION OF MOLECULAR MARKERS FOR CHARACTERIZATION OF LATVIAN CROP

PLANTS

Nils Rostoks University of Latvia

2012.05.31. Vienošanās Nr.

2009/0218/1DP/1.1.1.2.0/09/APIA/VIAA/099

2

Outline of the talk

• Genomics and plant breeding • DArT un SNP genotyping systems for

characterization of plant genomes • Trait mapping using association with SNP

markers • Markers for marker assisted selection

2012.05.31.2012.05.31. 2

3

Conventional plant breeding

2012.05.31. From Poehlman (1959) Breeding field crops 2012.05.31. 32012.05.31. 3

4

Genomics and plant breeding

2012.05.31.

Gen

omic

s

Plan

t bre

edin

g

Phenotype

Genetic map

Physical map

Genome sequence

Gene function

Allelic diversity

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DNA-based molecular markers

• Type of variation: - point mutations - insertions/deletions - repeat number variation - copy number variation

• Genotyping technology: - RFLP - AFLP - RAPD, ISSR - CAPS - SSAP, IRAP, REMAP - SSCP

2012.05.31.

Variation in anonymous genomic regions or variation in transcribed regions (genes)

Throughput and level of automation of the genotyping technology

6

DArT

• DArT genotyping available as service from the Diversity Arrays Technologies, Australia

• Dominant marker system, based on randomly digested polymorphic genomic DNA clones

• DArT platform available for barley, wheat, rice, oat and potato

• DArT genotyping done for Latvian barley, oat and potato accessions

2012.05.31.

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Oat DArT

2012.05.31.

-0.20 -0.15 -0.10 -0.05 0.00 0.05 0.10 0.15 0.20 0.25 0.30-0.15

-0.10

-0.05

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0.35

LV hullessLV hulledForeign hullessForeign hulled

PCo1 (18.7%)

PCo2

(9.2

%)

8

Oat DArT

2012.05.31.

0.1

31929Pob14391-93

3010431652

BOR88322

Bug

Hja82371

3129231995

3176932055

31960NSC-3076-76

D104

3170931029

EndspurtLaim

a3114231231

PCU-32

StendesDarta

3112431959Korol31611

Lia1313-137M

igla31143

30020Lia1313-100

3186932013

31945Belinda

FrejaKazstan31851

KirovecJum

boParsival

31796Zlatak

31824G

A931015IL85-2069

GA921098

P470731800Straļec

29427Skakun

P4690Panther

PennudaStendesVelas

StendesDzeltenas

32224StendesLiva

3135331759

ArtaH

echt3190131906D

101EP2S-156

31790Stm

araSW

9626931367

EditSelm

aBor-1426

Katri3219032191

Bor8860130629

29149Izak

OA

-504-5BelorusskijgolozernijA

damM

ernime28167

28168A

bel31868

3061630638

30636N

osNacht

2012.05.31. 82012.05.31. 8

92012.05.31.

Genomics for plant breeding

• Identification of genes encoding specific traits and characterization of gene function

• Assessment of allelic diversity in specific genes to link diversity and the gene function

• Markers for specific traits allowing to create varieties with defined properties

• Assessment of genetic diversity in germplasm available to plant breeders for new varieties with novel characteristics

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Development of molecular markers

- SNP discovery by resequencing- eSNP discovery from ESTs

Development of mapping populations

- Experimental populations - Natural populations

High-throughput genotyping Phenotyping

QTL mappingTrait mapping

Fine mapping

Development of low cost, robust markers linked to

specific traits

Chromosome walking/landing

Gene cloning Gene-based marker

development

Marker Assisted Selection

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Illumina SNP genotyping

• Genotyping technology developed by Illumina • Barley genotyping assays (BOPA1 and BOPA2)

developed by The James Hutton Institute, University of California, Riverside and The Leibniz Institute of Plant Genetics and Crop Plant Research

• 2 x 1536 SNP • Most of the SNPs mapped

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SNP diversity in Latvian barley germplasm

• 95 Latvian barley varieties and breeding lines genotyped with BOPA1 (1536 SNP)

• 96 Latvian and foreign accessions genotyped with BOPA1 and BOPA2 (2 x 1536 SNP)

• Assessement of genetic diversity based on BOPA1

• Association mapping in a set of 153 accessions genotyped with BOPA1

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SNP diversity in Latvian barley germplasm

2012.05.31.

0.1 Dzintars

VairogsL-2532

Priekulu1Agra

No-79

Daghestanicum

-PI330504N

o-511079488-45PR-3749

DruvisL24

ZNCF

PR-5108PR-5109

V-08-83Law

inaPR-5112

HeilsH

annaPR-5127

BZ14-90BZ14-99

BZ14-12PR-4803

Primus

3280-14-1-4PR-5105BZ12-86BZ12-93

69-Clho11319Kom

abainierisPR-4832LatvijasVietejie

PR-3274PR-3005PR-5131

1268498-3PR-3134

Danuta

PR-3885Tocada

PR-4822O

tiraPR-3512

PR-3518AnnabellJustina743-09

Heris

RoxanaSZD

4748N

uevo12811PR-4362PR-4368

PR-4115PR-4121PR-3421

PR-3473PR-3527

PR-3474PR-3475PR-3464

797877-39768678-28

827580-151163691-34

L-2630901683-48

RasaLeeni250-PI436150

AuraL-2529

L-2295PR-3245G

atePR-3528PR-3416PR-3414PR-3415

L-400PR-3467PR-3522

918683-13PR-3223SencisThuringiaH

ellanaSteffi

12820PR-3345

PR-3348Kristaps

ViennaBZ12-63

PR-3282EunovaPR-3335PR-3520

1272500-36754277-27

1273300-50PeggyPR-4760

1267199-30L-2544Cam

ilaVada12825

G-131

M9LysibaAnni

BetzesPervonez

L-2735AlsaVerena

718676-19PR-3300

PR-3515U

laPR-3807PR-3494

L13PR-3534M

aarenPR-3351

Inari1268799-2

L-2332M

ik1738177-41

Golf

PR-4181828680-15

1012786-41M

alva720076-34

742977-101271100-26

Klinta813380-13

StendesIlga

PallasPriekulu60

PR-3605PR-4814

B-93L-3008

PR-3487Ruja

12819803877-15

PR-5137PR-5135L-2406

AbavaAnsisD

ivosnojeIakub

Run8-458PR-3448

PR-3537PR-3438

PR-3440PR-3297

PR-3353L-3101

BalgaLinga

1263098-13PR-5145

BZ12-83Rubiola

PR-4810PR-4812

PR-4144PR-3636PR-4835

Bor88377PR-5117L-2233

L-2985 1H

130Im

ulaL14

Dziugiai

Idumeja

PCo2 (6.4%)

-0.2

-0.1

0

0.1

0.2

0.3

0.4

-0.4 -0.2 0 0.2 0.4

PCo1 (10.6%)

Foreign accession LV breding line

LV variety

LD (r

2)

0.10.20.30.40.50.60.70.80.9

1

0 50 100 150 200Distance between markers (cM)

Intrachromosomal Interchromosomal

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2H

3H

6H

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2009/0218/1DP/1.1.1.2.0/09/APIA/VIAA/099

Preliminary results of association mapping

• BOPA1 genotypes, phenotypes from biological and conventional trials 2010 – 2011

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b-glucan content

1 70 139 208 277 346 415 484 553 622 691 760 829 898 96710360

10

20

30

40

50

60

- Log

P

7H, 34.8 cM

1 70 139 208 277 346 415 484 553 622 691 760 829 898 96710360

1

2

3

4

5

6

7

8

- Log

P

Resistance to loose smut

6H, 55.7 cM

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Molecular markers and genetic diversity

• Genetic diversity is both the basis for development of varieties with different traits, and a tool to distinguish the varieties and map and transfer traits

• DNA-based markers directly assess genetic diversity in a genome

• Gene-based molecular markers allow to assess allelic diversity in genes providing direct link to functional diversity

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Markers for plant breeding

• Any marker that allows to select for a specific trait is useful in plant breeding

• Molecular markers detect differences between individuals or varieties at molecular (protein and DNA) level

• Molecular markers that are linked to certain traits form the basis for marker-assisted selection (MAS)

• Essentially, molecular markers are just another way of achieving what breeders have been doing for years – combine traits of interest in a novel variety in the most efficient way

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Markers for breeding

• Powdery mildew resistance in barley – Mla case

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Centromere ABC15612

~0.2 cM ~0.15 cM

538P8

RGH1bcd mla( )

Mla Telomere

Mla19 GB

Mla18 GB

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Outlook

• Association mapping in barley using SNP markers and phenotyping data collected within European Social Fund project

• Development of assays for published and new molecular markers for specific traits that can be used by breeders with a focus on traits important in organic farming

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Acknowledgements

• Plant molecular genetics group at the University of Latvia

• State Priekuli Plant Breeding Institute • State Stende Plant Breeding Institute • Funding – European Social Fund project

2009/0218/1DP/1.1.1.2.0/09/APIA/VIAA/099

2012.05.31.

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Acknowledgements

• Collaborators at the James Hutton Institute Dr. Robbie Waugh, Dr. William Thomas

• Collaborators at University of California, Riverside Dr. Timothy Close

• Diversity Arrays Technology Dr. Andrzej Kilian

2012.05.31.