Frédéric CHOULET A pseudomolecule of 774 Mb: the 3B experience - PowerPoint Presentation

Slide1

Frédéric

CHOULET

A pseudomolecule of 774 Mb: the 3B experience

INRA GDEC – Clermont-Ferrand, France

3B

Sequenced physical map



#

MTP

BACs

8452

3B MTP-BAC sequencing



#

BAC pools

922



#

Roche 8 kb MP lib.

922



bp

coverage

(Roche/454)

36x



BAC-ends

(Sanger)

42,551



Whole Genome Prof. tags

327,282



Whole 3B shotgun

(Illumina)

82x



#

BACs



#BAC-

contigs

Physical map

132,000 (19x)

1282



#

MTP-BACs

8452

3B

900 Mb

3B physical map

ACGTAGACTACA

3B

Assembly and scaffolding

3B-v1

16,136

scaff

1,040

Mb

Curation of the scaffolding

V. Barbe, S.

Mangenot (Genoscope)

Assembly and scaffolding

Integration of

BAC-end

match positions

Parsing of MP read positions

scaff00001

scaff00013

scaff00024

scaff00008

scaff00011

scaff00007

scaff00005

3B-v1

16,136

scaff

1,040

Mb

18% Ns

3B-v3

4,999

scaff

992

Mb

13% Ns

Curation of the scaffolding

V.

Barbe, S. Mangenot

(

Genoscope

)

Assembly and scaffolding

3B-v3

4,999

scaff

992

Mb

1

3% Ns

3B-v1

16,136

scaff

1,040 Mb

Gap filling

Seq. error corrections

JM.

Aury

, A.

Couloux

(

Genoscope

)

Illumina

reads

Whole

3B Shotgun

109,914 gaps filled

126,290 bases corrected

(error rate: 0.1%)

18% Ns

3B-v4

-

-

8

% Ns

Curation of the scaffolding

V.

Barbe, S. Mangenot

(

Genoscope

)

Assembly and scaffolding

3B-v1

16,136

scaff

1,040

Mb

Gap filling

Seq. error corrections

JM.

Aury

, A.

Couloux

(

Genoscope)

Redundancy removal and scaffold merging

S.

Theil (INRA GDEC)

Pool_A

Pool_B

ctg1

ctg2

2,808

scaff

833 Mb

3B-v443

scaffAssembler.pl

3B-v4

4,999

scaff

992

Mb

redundancy:

160 Mb

Search for shared TE-junctions

Ordering scaffolds

2,808

scaff

833 Mb

3B-v443

SNP discovery

Bait

TE

DNA captured from 10 genotypes

gene

52,265 baits

isbpProbeDesign.pl

39,077 SNPs

SureSelect

®

seq. capture

(

E.

Paux

, N.

Cubizolles

, E. Rey

)

Ordering scaffolds

SNP discovery

Genetic mapping

(

P.

Sourdille

)

+ Neighbor map: 3865 markers

LD mapping

(

F.

Balfourier

)Anchor map: 384 indiv

Cs x Renan Genotyping mapping pop

3,075 SNPs367 lines from a core-

collection

3B

Ordering scaffolds

genetic map

4

4.8

cM

152 scaffolds

LD map

19

LD blocks

366 bins

0

cM

133

cM

554 bins

64 markers

at

the same genetic position

Linkage Disequilibrium

Ordering scaffolds

pseudomolBuilder

.pl

1358

scaff

774 Mb

pseudomolecule

unlocalized

1450

scaff

59 Mb

93%

7%

N

N

N

N

N

N

N

N

SNP discovery

Genotyping mapping pop

Integration of phys. map info

3

cM

0

1

2

3

3

4

5

6

A

B

C

D

E

orientation unknown:

48%

of the seq.

micro-order unknown:

554 bins / 1358

scaff

?

?

RH map

Optical map

Long reads

Future Improvements

?

7264

protein

coding genes

TRIANNOT

234,606

TEs

CLARI-TE

774 Mb

Annotation

Bioinformatics

Scaffolding/pseudomolecule construction

scaffAssembler.pl

Annotation

gapCloser

ssrFinishing

triAnnot

(new modules: filtering, pseudogenes, transfer annotation)

clari

-TE &

clari

-TE-lib

Data management

gowDB

(Bio::DB::

seqFeatureStore

)

Gbrowse

@ URGI

pseudomolBuilder

.pl

isbpProbeDesign.pl

Assembly

Newbler

3B-MTP

3B-CSS

Pseudomolecule

vs

GenomeZipper

Sébastien

Theil

Natasha Glover

Josquin Daron

Lise

Pingault

Pierre Sourdille

Etienne Paux

Philippe Leroy

Jacques Le

Gouis

Nicolas Guilhot

Aurélien

Bernard

Nelly Cubizolles

Catherine Feuillet

François

Balfourier

M.

Alaux

L.

Couderc

V.

Jamilloux

H.

Quenesville

URGI

H. Berges

A. Bellec

CNRGV

BIA

A.

AlbertiV.

BarbeJ. Poulain

C. DurandS.

Mangenot

JM. Aury

A. Couloux

P. Wincker

Genoscope

J.

Dolezel

J. Safar

IEB

K.

Vandepoele

K. Mayer et al.

P.

Schnable

S.

Rounsley

D. Ware

C.

Gaspin

SAB

VIB

MIPS

Acknowledgments

Hélène

Rimbert

TGAC

J.

Rogers, M.

Caccamo

et al.

J. Rogers

K.

Eversole