Parkin et al. 2005 (Genetics) 1 Collinearity defined: - mosaic of segmental blocks Brassica A + C genomes: B. napus 234 5 A B C D E A B B C Arabidopsis.

Parkin et al. 2005 (Genetics)

1

Collinearity defined: - mosaic of segmental blocks

Brassica A + C genomes: B. napus

2 3 4 5

A

B

C

D

E

A

B

B

C

Arabidopsis

A

B

C

D

E

A

B

C

E

Database developments

• adopt GERMINATE as public-domain repository

• curate data locally in CropStore

• use agreed nomenclature

• input template spreadsheets

• parse into GERMINATE

• Perl-cgi Web interface

Fady Mohareb: MSc project at RRES with Cranfield

OREGIN data presented from GERMINATE interface

Populations (TN, DY, et al)BnDFS, BnDFFSMaps (TN, DY, Parkin et al.)Marker loci and sequence-tags

Links to A and C genome maps

Brassica linkage maps, in GERMINATE with CMAP (GMOD) viewer

Loci will link to genomic resources

CKDH sensu Parkin et al., Sharpe et al.

NIAB LGsNIAB

chromosomesNVITS, Japan (Suwabe) LGs

Chiifu (CNU) chromosomes sensu Koo et al

CNU CK LGs

R1 - R10 = N1 - N10

Kim et al. Lim et al. Choi et al.

R1 LG10 Chr8 6   LG9

R2 LG3 Chr6 8   LG6

R3 LG1 Chr2 1   LG7

R4 LG4 Chr3 10   LG2

R5 LG7 Chr5 3   LG8

R6 LG5 Chr4 2   LG1

R7 LG6 Chr7 4 ?KooR2 LG5

R8 LG9 Chr9 7 ?KooR9 LG10

R9 LG2 Chr1 5 KooR1 LG4

R10 LG8 Chr10 9   LG3

Rosetta Stone for Rosetta Stone for BrassicaBrassica linkage group/chromosome assignments linkage group/chromosome assignments

Parkin et al., (2005) map based on 1000 RFLP markers

Kim et al., (2005) map based on >500 RFLP markers

CK reference B. rapa map paper almost ready for submission

Ol12F110.0

BRMS-05633.6

RA2E0450.0

BRMS-02473.8

Na14D07121.1

N01

BRMS-2151.1

Na10B1018.0

BRMS-02656.1

Ol10F0480.0

Ol10A0590.6

N02

BRMS-04315.8

BRMS-10631.9

BRMS-04269.4

Na12A0190.6

BRMS-008108.0

Ol10B01232.0

N03

BRMS-00113.3

BRMS-05426.7

BRMS-19541.5

Na12A0190.6

N04

BRMS-03413.0BRMS-06120.8

BRMS-24252.4

BRMS-00780.2

Ra3H0193.0

N05

Na12B080.0

Na12H0722.0

Ra1F0648.2BRMS-02755.0

Na12D04134.8

N06

RA2G081.0

BRMS-01822.0

Ra2A0158.5

BRMS-03669.9

Ra2A0592.6

N07

BRMS-0940.0

Ra2E1220.5

BRMS-08831.8

BRMS-24642.6

BRMS-00683.4

N08

Ni4D0929.5

BRMS-02954.9Ra2A1164.4

BRMS-24780.2

BRMS-06096.3

N09

Ol11B030.0

Na10D0745.0Ra2E0746.6BRMS-01956.9Ni4A0358.9

N10

REM1b-SSR33.0

Nga24848.8

Ol10F1166.0Na12C0872.9

Ni4B10108.0

N11

Ol12B0336.0Ni2C1242.6

Ol13G0575.0

Na12C0389.3

Ol11H09105.0

N12

Na10G100.0

Ra2E1220.5

Na14E0254.4

CALSSR(LSI07)94.0BN12A98.0

Na12F12115.2

N13

Ol10F1226.1

Ni4F0849.3

Na14E0876.1

Ol12D0292.0Na12E05102.1

N14

Ol10A105.0

Ni4C1138.1Na12D1043.9

sORA2166.6

Na12G1295.0

N15

Ol10F0916.9

NGA11129.7

Ca7264.4

BRMS-01593.0Na12A05101.0

N16

Na12F036.1

CHIA-SSR40.5

BN35D63.5

BN72A80.8

Ol10B01232.0

N17

BN35D7.8

Ol12G0447.6

Ol12D0562.1MT1-SSR69.1

N18

Ol11B030.0Ni2B0110.1

Ol12A0428.9

Na10D0745.0Ol13C0351.9

BN83B163.8

Ol11H0689.9

N19

Selection of markers for diversity screeningSelection of markers for diversity screening

Reactions performed on subset 1 with 12 markers highlighted in pink

Analysis will be performed using BioGene GeneMarker software to replace ABI GeneMapper

Testing fidelity of GenomiPhiTesting fidelity of GenomiPhiTMTM amplification amplification

Line name Source Na12A07 Na10A08

Marinka NGB 5 3

Niklas BAZ 4 3

Two lines showing high allelic diversity with two SSR markers have been identified

Experimental design:

• Genomic preps performed from 8 plants each of Marinka and Niklas• 4 GenomiPhi amplifications performed for each genomic DNA prep• These will each be assayed with the above two SSRs and compared with duplicate SSR

reactions performed on the original genomic DNA

The control will be 3 genomic preps from Tapidor (DH) each amplified 4 times with GenomiPhi

• We are planning to perform our diversity screening on, and to distrtibute genomic DNA amplified with GenomiPhi.

• Concerns regarding the fidelity of GenomiPhi were raised at the last OREGIN Stekeholder meeting

• So we need to estimate the likely mutation rate resulting from the GenomiPhi reactions

No. of alleles detected in a sample of 8 plants per line:

Acquisition of Tapidor x Victor substitution linesAcquisition of Tapidor x Victor substitution lines

Graham King has requested permission to adopt this population within OREGIN:

Peter Werner, CPB Twyfords – yes

Steve Barns, Advanta, subsequently sold to Limagrain – yes

Mike Kearsey, Birmingham – yes

Jo Bowman, Nickersons – yes

Monsanto – awaiting reply

TN PopulationTN Population

2005 field bulking at WHRI of core set of 50 lines

For QTL analysis really want 100 lines bulked - RRES may sow additional 75 lines

Remainder of TN population is being regenerated in the GH

Harvesting in progress

BnaDFFS Microspore cultureBnaDFFS Microspore culture

Microspore culture progressMicrospore culture progress

Line Name No. of plants produced

4n DH 1n Colchicine doubling

No. awaiting ploidy analysis

Plants still in jars

Plants still in plates

Fido 41 2 26 3   8   2

Duplo 60   22 19   23 15 11

Couve nabica 41 18 11 8 1 3 5 3

Emerald 9 2 3     3 4  

Cubs Root 6   2     4    

Global 11   2 1   8 2 7

Matador 1   1     0    

Apex 1   1     0 2 3

Judzae 8   1 5   2    

Jet Neuf 2   1     1 1 3

Abukuma Natane 8   1 2 4 3   3

Hanna 3     2   1 3 1

Guelzower 5     1   4   3

Brutor 3     1   2 4 4

Bronowski 4 1   1   2    

Ceska 1         1 4 4

Canard 7     6 6 1 4 16

Sarapta               128

Moana Moana Rape               182

Dippes               130

Brauner Schnitkohl               21

Line Colchicine - yes   Colchicine - no  

  1n 2n 4n 1n 2n 4n

Apex   1        

Bronowski     1 1    

Brutor       1    

Couve nabica   10 18 8 1  

Cubs Root   2        

Duplo 5 18   14 1  

Emerald     2   2  

Fido   22 2 3 3  

Global   2   1    

Guelzower       1    

Hanna       2    

Jet Neuf         1  

Judzae 1 1   4    

Matador         1  

Effects of colchicine treatment in the initial mediumEffects of colchicine treatment in the initial medium

Bulking of the founder linesBulking of the founder lines

Founder lines being regenerated with a target of >10g seed per line at WHRI

In addition:

29 founder lines sent to Mark Nightingale, Elsoms for bulking this 2005/06• Will bag 2 plants/line – 25-100g seed for their seed• Collect unbagged seed from middle of 1.2 x 3 m plot – upto 1 kg• Seed yields expected to be lower for DFS lines

36 fixed lines sent to Neal Evans, RRES, to include in their 2005/06 field bulking

Possibility of distributing of seed through NASC - need to discuss further with Sean May

MTA?MTA?

What next for OREGIN?What next for OREGIN?

Some thoughts from WHRI

Plant resourcesWant to maximise ability to utilise the BnaDFFS and mapping populations• Complete fixation of the BnaDFFS• Bulk up BnaDFFS, TN, DY mapping pops and TV substitution lines - breeders?• Possibly extend BnaDFFS in association with Chinese

Marker analysis• Complete genotyping of DFFS with 100 SSRs• AFLP fingerprint DFFS – provides alternative diversity assessment and quick QC check

– fairly breeder friendly• Generate sufficient marker density to enable association analysis (>1000) - other

funding sources for this• EcoTILLING genes of interest

Pathogens• UK core collection from existing collection

Trait analysis• Oil analysis• Screen pathogen core collection against BnaDFFS• Others, e.g flowering time, water use efficiency, mineral use efficiency, virus

resistance, vernalisaiton requirement, other seed traits

Data management• To include collation of published QTLs