The VIGNA Project: Genomic PlatformsImplementation in Italy

The French-Italian Public Consortium for theSequencing of the Grapevine Nuclear Genome

M. Enrico PèScuola Superiore di Studi Universitari e Perfezionamento

Sant’Anna di Pisa

RESEARCH PHYLOSOPHY

Genomic Project with Grape as the Object Organism of Choice

Genomic Goals

To Implement National PlatformsFor Genomic Analysis

a) technological upgradeb) formation of a new generation

of young scientists with genomic skills

Applied Research

To Develop and ApplyGenomic Tools to grape

a) Applied Pilot Projectsb) Interface with specialists and researchers in the

field

Sequencing MappingPhysical map

High resolution genetic mapQTLs for resistance

LEVEL 1 Structural Genomics

Germplasm characterization

Berrymaturation

LEVEL 3

Bioticstresses

Abioticstresses

Applied Projects

ReproductionDevelopment

Transcriptomics BioinformaticsEST

MicroarrayWeb portalDatabase

Data analysis

LEVEL 2

Proteomics Metabolomics2D-EMS

Functional Genomics

PROJECT STRUCTURE

ACTIVITY STARTED AT ALL LEVELS AND IN ALL GROUPSin JANUARY 2006

Consorzio Interuniversitario Nazionaleper la Biologia Molecolare delle Piante CINBMP

11 research groups

ENEA Centro Ricerche Casaccia, Rome – Eugenio BenvenutoCNR Dip. di Bioinformatica e Genomica, Bari – Sabino Liuni e Cecila SacconeCIB Consorzio Interuniversitario per le Biotecnologie, Trieste – Claudio SchneiderUniversità di Bari Dip. di Biologia Molecolare – Graziano PesoleUniversità di Milano Dip. Scienze biomolecolari e Biotecnologie – M. Enrico PèUniversità di Padova CRIBI Centro per le Biotecnologie – Giorgio ValleUniversità di Siena Dip. di Scienze Ambientali Sciences– Mauro CrestiUniversità di Udine1 Dip. Scienze Agrarie e Ambientali – Michele MorganteUniversità di Udine2 Dip. Scienze Agrarie e Ambientali – Raffaele TestolinUniversità di Verona1 Dip. di Scienze – Massimo DelledonneUniversità di Verona2 Dip. Scienze, Tecnol. e Mercato della vite e del vino – Mario Pezzotti

CRA Istituto Sperimentale per la Cerealicoltura di Fiorenzuola d’ArdaPlant/Pathogen Interaction – Luigi CattivelliPhytopatology – Giampiero ValèBioinformatics – Primetta FaccioliCRA Istituto Sperimentale per la Viticoltura di Conegliano venetoGermplasm characterization - Antonio CalòSugars Metabolism – Angelo CostacurtaPlant Development - Manna Crespan

Consiglio per la Ricerca in Agricoltura CRA6 research groups

Coordinator: Michele Stanca

GRAPE GENOME SEQUENCING~480 million bp

FINAL GOAL: 12 GENOMES EQUIVALENTS (12x)

Using PN40024 homozygous clone (~ 93%)derived from Pinot noir and Helfesteiner after 6 cycles

of selfing (Colmar, Fr)

ITALY5 x

FRANCEGENOSCOPE

7 x

CRIBIPadova2.0 x

IGAUdine3.0 x

Sequencing activity started January 2006Completed August 2007

Progress since April 2007

• 12 x coverage of PN40024

• 90% sequences organized in < 150 supercontigs

• > 90% sequences anchored to genetic maps

• 0.9 x coverage of cultivar Friulano

• 1 SNP every 98 bp identified

• 1.3 million SNPs estimated

In Progress• Deep cDNA sequencing by Solexa/Illumina and 454

technology for annotation improvement

• DNA arrays using Combimatrix technology for

Complete transcriptomics analysis

Accomplished

Arachne 8.4X

Pcap 12X

Pairedsupercontig(consensus)

Assembling assemblies:positioning supercontigs into a continuous

assembly

Preliminary consensus: whole genome in~150 ultracontigs

Contig 12 Contig427Contig 68

Contig BContig A

Strategy for detecting structural variation

Friulano < PN40024:Insertion in PN

Friulano > PN40024:Insertion in Friulano

Friulano paired reads

Friulano paired reads

Deep cDNA Sequencing for ImplementingGene Prediction and Genome Annotation

1. New sequencing technologies produce millions ofshort DNA sequences

2. Utilized for sequencing of expressed genesa) from berries at different stage of maturation

350 thousands 250 nucleotide long readsb) from infected tissues

200 thousands 250 nucleotide long readsc) from leaves

40 millions 35 nucleotide long reads

Other tissues in progress

20 – 22 nucleotide long MicroRNAs areImportant Regulatory Elements Coded by

MIR Genes as Precursors

Validation of MIR gene prediction by deepcDNA sequencing

TRAINING ACTIVITY

A Major Objective of the VIGNA

43 YOUNG SCIENTISTS CURRENTLY

INOLVED IN THE DIFFERENT VIGNA

WORKPACKAGES

(37 ON VIGNA GRANT)

VIGNA FINACIAL RESOURCES

• MiPAF: € 6.5 millionswithin the French – Italian bilateral agreementsigned by INRA and CRA

• Instituto di Genomica Applicata, Udine € 2 millions

• Regione Veneto, Indistrial Sector€ 1.5 milioni

• Bank FoundationsCariverona € 0.50 millionMonte dei Paschi di Siena € 0.17 million

From the Grape Genome GreatDiscoveries and Surprises

The French-Italian Consortium

Olivier JaillonJean-Marc AuryFrancis QuetierPatrick WinckerJean Weissenbachet al.

Michele Morgante, Alberto PolicritiGabriele De Gaspero, Federica Cattonaro,Alberto Casagrande, Cristian Del Fabbro,Nicoletta Felice, Irena Iurman, SimoneScalabrin,

Anne-Francoise Adam-BlondonChristian ClepetMichel CabocheSebastien AubourgFabrice LegeaiDidier Merdinogluet al.

CRIBI Padova UniMiGiorgio Valle David HornerNicola Vitulo Erica MicaAlessandro Vezzi M. Enrico PèGiorgio Malacrida

UniBa UniVrGraziano Pesole Mario Pezzotti

Massimo Delledonne