Durum wheat genome highlights past domestication ...Ta cv. Chinese Spring Illumina/PacBio reads...

Durum wheat genome highlights past domesticationsignatures and future improvement targets

Gabriella Sonnante

Istituto di Bioscienze e Biorisorse - CNR

Conferenza DiSBA – CNR 12-13 dicembre 2019

Durum wheat (or “pasta” wheat) is an important food crop in

the world, with an estimated annual global production of 36

million t.

Turkey and Canada are the largest producing countries, with

estimated 2 million ha each, followed by Algeria, Italy and

India, each cultivating over 1.5 million ha. Before war, Syria

belonged to this group of large producers.

A very large amount of genetic diversity exists in durum

wheat germplasm, and that diversity also extends to the

many traditional ways of consuming durum, including

special dishes that represent the national identities for

some Mediterranean countries: pasta, couscous,

bourghul, freekeh, gofio, particular breads, etc.

Durum wheat (Triticum turgidum ssp. durum)

Human driven teraploid wheat evolution

WILD EMMER WHEAT

Domestication

DOMESTICATED EMMER WHEAT

Continued evolution under Domestication

DURUM WHEAT LANDRACES DOMESTICATED EMMER WHEAT

MODERN DURUM WHEAT CULTIVARS

Breeding improvement

DURUM WHEAT LANDRACES

- Fully assembled genome of the modern cv. Svevo

- Genome-wide account of modifications imposed by thousands of years of empirical selection and breeding

- comparing the Svevo genome with the assembled genome of Wild Emmer Wheat accession Zavitan

- through a survey of the genetic diversity and selection signatures in a Global Tetraploid Wheat Collection

consisting of 1,856 accessions

- Efficient, genome-enabled dissection of the Cdu-B1 locus. A region bearing a signature of historic selection co-locates

with Cdu-B1, a quantitative trait locus (QTL) spanning 0.7 cM on chromosome 5B known to control cadmium

(Cd) accumulation in the grain

Durum wheat genome

# Library type Insert sizeSequencing

InstrumentRead length

Minimal

Coverage

1 PCR-free paired end library 450-460bp HiSeq 2500 PE 250bp X123

2 PCR-free paired end library 700- 800bp HiSeqX PE 150-160bp X38

3 Mate pair library 2-4kbp HiSeqX PE 150-160bp X39



Detailed sequencing data

Sequencing output

3.35 terabases (Tb) of data

279 × coverage (estimated genome size of 12 Gb)

Contigs Scaffolds

Total number of sequences 474,837 129,464

Assembly size (bp) 10,299,203,836 10,450,113,204

Gaps size (bp) 0 149,190,797

Gaps % 0.00 1.42

N50-length (bp) 56,196 5,972,063

N50-number of sequences 54,378 493

N90-length (bp) 13,008 1,085,649

N90-number of sequences 194,738 2,019

Maximal length (bp) 475,246 54,262,061

Detailed assembly results – (DenovoMAGIC2-NRGene)

Chromosome Sequence length Number of scaffolds

chr1A 585,266,722 126

chr1B 681,112,512 248

chr2A 775,448,786 167

chr2B 790,338,525 247

chr3A 746,673,839 190

chr3B 836,514,780 275

chr4A 736,872,137 241

chr4B 676,292,951 197

chr5A 669,155,517 155

chr5B 701,372,996 247

chr6A 615,672,275 140

chr6B 698,614,761 257

chr7A 728,031,845 198

chr7B 722,970,987 250

chrUn 498,719,471 126,526

Summary statistics of the durum wheat pseudomolecules (two

Hi-C libraries)

Scaffolds ordered and oriented using Svevo x Zavitan genetic map.

Conformation capture sequencing (Hi-C) resulted in 14

pseudomolecules (chromosomes) and a group of unassigned

scaffolds.

Pseudomolecules encompass 93.5% of the assembled sequence,

90% of the scaffolds oriented

Material File in public repository Tissue Reference

T. durum (Td) cv. Svevo This study 57 treatments seedling /adult plants - 9 RNA Pools This study

Td cv. Svevo This study Grains at 5 levels of developments This study

Td cv. Senatore Cappelli This study Grains at 5 levels of developments This study

Ta cv. Chinese Spring Illumina/PacBio reads PRJEB15048 Leaf; Root; Seedling; Seed; Stem; Spike Clavijo et al, 2016

Td cv. Kronos Bioproject PRJNA191054 young roots; young shoots; spike; grain Krasileva et al. 2013

Ta cv. Chinese Spring RNA-Seq (acc. number ERP004714) grain; leaf; root; spike; stem Pingault et al. 2015

Td cv. Altar84; Capeiti8; Claudio; Cresco; Edmore; Kofa; Meridiano;

Neodur; Saragolla; Strongfield; ValnovaThis study anther+ovary; root; leaf

Td cv. Svevo This study anther+ovary; root; leaf; seed_anthesis; seed_milk

T. turgidum dicoccoides “Zavitan” accession WEW: BioProject PRJNA310175Leaf; root; flag leaf; developing spikes; glumes;

flowers; grainsAvni et al. 2017

Two wild emmer, two landraces, two durum cultivars SRA (acc. N: SRR2084071 etc.) Glumes Zou et al 2015

RNA seq

Annotation

- Genes/Proteins (High Confidence; Low Confidence)- Gene Functions- Repeat Elements- microRNAs (from sRNA libraries)- Long non-coding RNAs- NLR gene family (Nucleotide Leucin-Rich Repeat Domain)- CpG islands- Prolamin proteins (Glutenins, Gliadins, Avenin-like)- Identification of pseudogenes- Orthologous gene family analysis- Genome-wide atlas of putative functional variants (SNPs-indels) between WEW and Durum wheat. In total, 597 variants.

Svevo (DW) – Zavitan (WEW) – Strong overall synteny

66,559 – 67,182 – Total High Confidence gene number

Similar chromosome structure and Transposable Element composition

Structural, functional and conserved synteny landscape of

the DW genome

a - Chromosome name and size (arms differentiated by gray shading)

b - Density of WEW High Confidence (HC) gene models

c - Links connecting homologous genes between WEW and DW

d - Density of DW HC gene models

e - Location of published QTLs

f - k-mer frequencies

g - Long terminal repeat (LTR)-retrotransposon density

h - DNA transposon frequency

i - Mean expression of HC genes calculated

Links in center connect homoeologous genes between subgenomes;

blue links between homoeologous chromosomes and

green links between large translocated regions.

Germplasm structure and phylogenetic relationships

Global Tetraploid Wheat Collection

- 1,856 accessions (non-redundant, filtered. Originally 2,558)

four main germplasm groups involved in tetraploid wheat domestication history and breeding:

Wild Emmer Wheat (WEW), Domesticated Emmer Wheat (DEW),

Durum Wheat Landraces (DWL), Durum Wheat Cultivars (DWC)

- 17,340 SNPs (90K SNP Infinium assay - unique, non –redundant, single Mendelian markers, genetically and

physically mapped) used for:

- Genetic Diversity

- Population Structure

- Identification of Selection Signature

DEW

DEW-ETH

DWL-ETH

DWLDWL DWL

DWL

DWC

DWCDEW

WEW-SL

DWL

DWL-ETH

DEW-ETH

DWC

WEW-SL

WEW-NE

Global Tetraploid Wheat Collection – Population Structure

Admixture analysis

Wild Emmer Wheat

WEW from North-Eastern Fertile

Crescent (WEW-NE)

Turkey-Karakadag pop 1




Iran-NW (Kermanshaha) / Iraq

Admixed accession

WEW from Southern Levant

Fertile Crescent (WEW-SL)

Israel Northern pop 1

Israel Northern pop 2

Israel central pop 3

Jordan

Syria

Turkey-Gaziantep (admixed NE-SL)

Lebanon

k=2*

k=3

k=4

k=5

k=6

k=7

k=8

k=9

k=10

k=11

k=12

Domesticated Emmer Wheat DEW

populations (K=6*)

Turkey to TransCaucasia

Turkey to Balkans

Southern Europe

Southern Levant to Europe 1

Southern Levant to Europe 2

India-Oman-Ethiopia

k=2

k=3

k=4

k=5

k=6*

k=7

k=8

k=9

k=10

k=11

k=12

k=13

k=14

k=15

k=16

k=17

k=18

k=19

k=20

Durum Wheat Landraces

Southern Levant to North Africa

Greece to Balkans

Turkey to Trans Caucasia

Turkey to whole Fertile Crescent

T. turanicum subpopulation

India-Oman-Ethiopia

k=2

k=3

k=4

k=5

k=6*

k=7

k=8

k=9

k=10

k=11

k=12

Durum Wheat Cultivars DWC populations (K=5*)

ICARDA and Italian breeding for dry-land areas

Core Italian breeding

Core CIMMYT-70 wide adaptation and ICARDA

breeding for temperate areas

Core CIMMYT-80 high yielding potential

North American-French breeding

k=2

k=3

k=4

k=5*

Average rate of

diversity reduction

from WEW to DWC:

32.6% (SNP-based),

espec. In peri-

centromeric

recombination depleted region

SNP-based

diversity index

for the main

germplasm

groups in the

Global

Tetraploid

Wheat Collection

DRI: Diversity reduction index Fst: Single site divergence index XP-EHH: Cross population haplotype homozygosity

XP-CLR: Multilocus test for allele frequency differentiation hapFLK: Haplotype-based differentiation test

Total 454 selection signals identified by at least 1 metrics Cross-

population

selection index

metrics for the

comparison between DWL

and DWC

VRN-A1

tough-glumesQTL

DRI: Diversity reduction index Fst: Single site divergence index XP-EHH: Cross population haplotype homozygosity

XP-CLR: Multilocus test for allele frequency differentiation hapFLK: Haplotype-based differentiation test

Cross-

population

selection index

metrics for the

comparison between DWL

and DWC

Cross-

population

selection index

metrics for the

comparison

between DEWand DWL

Disease resistanceYellow pigment (Psy-1)

Glu-A1

Grain proteincontent QTLsWEW to DEWDEW to DWL

a) The Cdu-B1 locus was mapped in three bi- parentalpopulations (G9586, 8962-TL and Svevo × Zavitan) to beon chromosome 5B [QTL accounts >80% of thephenotypic variation in Cd concentration].

b) High-throughput sequencing of exomes refined theCdu-B1 locus by identifying additional polymorphismassociated with low (blue) and high (black) Cdaccumulation; positions of markers from the originalmapping populations are indicated

c) Alignment of Cdu-B1 between Svevo and Zavitanidentified a region of increased sequence variation. Thisregion contains a variable gene that encodes a metaltransporter, HMA3-B1 (blue)

Functional analyses demonstrated that the alleleTdHMA3-B1b (but not TdHMA3-B1a) is responsible forCd accumulation

4.3 Mbp Fine Mapping (8982-TL)

7.2 Mbp Weibe et al. 2010 (G9586)

Genome Alignment (Svevo vs. Zavitan)

Cdu-B1 Variable Region

High Cd

Haplotypes

Low Cd

Haplotypes

90k (G9586)

90k (Svevo x Zavitan)

SNPs and

InDels

per kbp

Genes

Using the Svevo genome to identify a candidate gene associated with differences in cadmium (Cd) accumulation in the grain of durum wheat

TdHMA3-B1a

TdHMA3-B1b

https://www.interomics.eu/durum-wheat-genome

https://www.interomics.eu/durum-wheat-genome-intranet

https://www.interomics.eu/durum-wheat-genome

https://www.interomics.eu/durum-wheat-genome-intranet

D. Nigro

A. Gadaleta

University of

Bari, Italy

R. Tuberosa

M. Maccaferri

D. Ormanbekova

E. Frascaroli

S. Corneti

S. Salvi

University of

Bologna, Italy

H. Budak

S. Biyiklioglu

Montana

State

University

USA

A.T.O. Melo

I. Hale

University

New

Hampshire

USA

K.F.X. Mayer

S.O. Twardziok

H. Gundlach

M. Spannagl

T. Lux

V.M. Prade

Helmholtz

Zentrum

München,

Germany

B. Kilian Global Crop

Diversity Trust,

Germany

N. Stein

S.G. Milner

M. Mascher

A. Himmelbach

Leibniz Institute

Plant Genetics,

Germany

N. Pecchioni

P. De Vita

D. Marone

A.M. Mastrangelo

CREA

Foggia, Italy

L. Cattivelli

E. Mica

C. Marè

E. Mazzucotelli

P. Bagnaresi

P. Faccioli

F. Desiderio

C. Crosatti

CREA

Fiorenzuola

d’Arda, Italy

S.S. Xu

S. Chao

J.D. Faris

E.T. Schafer

Agricult. Res.

Center, USA

Washington

State

University

USA

M. Pumphery

L. Milanesi

A. Manconi

M. Gnocchi

M. Moscatelli

CNR-ITB

Segrate, Italy

CNR-IBBA

Milano, Italy

A. Ceriotti

A. Stella

P. Cozzi

M. Lauria

B. Lazzari

G. Sonnante CNR-IBBR

Bari, Italy

A. Distelfed

R. Avni

J. Deek

Tel Aviv

University,

Israel

R.K. Pasam

R. Joukhadar

M.J. Hayden

Agriculture

Victoria,

Australia

C.J. Pozniak

K. Wiebe

J. Ens

R.P. MacLachlan

J.M. Clarke

S. Walkowiak

A. Sharpe

C. S. Koh

University of

Saskatchewan,

Canada

R. Knox Swift Current Res.

Develop. Centre

Canada

N. S. Harris

K.Y.H. Liang

G.J. Taylor

University

of Alberta,

Canada

H. Özkan Çukurova

University,

Turkey

Durum wheat genome highlights past domestication ...Ta cv. Chinese Spring Illumina/PacBio reads...

Documents

Transcript of Durum wheat genome highlights past domestication ...Ta cv. Chinese Spring Illumina/PacBio reads...