Chr9

Antonio GranellIBMCP-Valencia

Spain

Tomato Sequencing, Madison July 2006

-Most markers are in heterochromatin

-Most of them did not match any BAC

-Gap of 46cM

Chromosome 9 has a total of 142 markers

But...

264 BACs from the tomato HindIII library were obtained from Cornell on July 2005. We started with a number of “seed BACs” following the recommendations

of estimated size and resent in a contig from FPC maps.

Selection of best seed BACs (>100Kb, in a contig, with known interest for scientific community)

New selection of candidates as “seed BAC”. Screening of 16 markers. 2-3

different BAC clones screened for each marker

These BACs will be used as seed BAC, if marker is

found, despite risk

Sequencing

Selection of BACs to extend

6 BAC clones as candidates for “seed

BACs”

Screening of markers

Just 2 BACs contained marker

Some did not have the marker, others contained the marker but were not in a contig or did not have an estimation of their length

10 “seed BACs” although some had a length of <100Kb

New screening of markers

SO FAR

BACs at different progress levels

5 confirmed, 13 possible.Several problems.

4 seed BACs more. Total of 14

Location in Chr9 for seed BACs done using Dani’s lines and FISH done by Song-Bin Chang and Steve Stack

Le_HBa0026I24

Le_HBa0116C14 14

23 Le_HBa0203J14

Le_HBa0168F14 37.5

46

48Le_HBa0026P14Le_HBa0300E15

H

* Completed BACs

Seed BACs

* BACs in progress* Candidates Extending BACs

Extending BACs

Currrent status of chromosome 9

Short arm

SL_MobI0144H04

Le_HBa0148A06/SL_EcoRI0001L13

SL_EcoRI0130H12

SL_EcoRI0019J03

SL_EcoRI0032F16

SL_EcoRI0004H20

SL_EcoRI0022M12SL_EcoRI0103M07/ SL_EcoRI0116G11

SL_EcoRI0004D19

Le_HBa0107D15

Le_HBa0099F1457

60 Le_HBa0099P03

112 Le_HBa0109D11

Le_HBa0278J12 61

Le_HBa0183J02116

Le_HBa0226D21 109

Le_HBa0165P17107

H

Currrent status of chromosome 9

Long arm

* Completed BACs

Seed BACs

* BACs in progress* Candidates extending BACs

Extending BACs

Le_HBa0059I05

Le_HBa0033H16

Le_HBa0015O02

Le_HBa0234F21Le_HBa0008K01

SL_EcoRI0049P19

Le_HBa0226J12

SL_MboI0017K18

Le_HBa0099F14SL_EcoRI0079M11 ! SL_EcoRI0079M11

No matches found when searching in tomato BAC end database. Too large overlaps (>20 kb, ex. SL_EcoRI0004D19 or Le_HBa0038L16). Some requested extending clones were contaminated with other clones

(ex. SL_EcoRI0004H20 contaminated with SL_EcoRI0004H19) or just were not in the sample sent (ex. Le_HBa0015O02, requested twice, both times was a mix of Le_Hba0014O02 and other clones but Le_HBa0015O02).

Estimated size BAC inserts far from actual size (ex. Le_HBa0033H16) For some clones, no restriction digest data available to check size of the

BAC although an estimation of its length is given (ex. Le_Hba0033H16)

Problems when extending finished BACs

Alternatives:•Selection of new seed BACs in order to keep working waiting for FPC contig map to be completed with new libraries.•Go for clones from new libraries that neither have an estimation of their length nor are present in FPC contigs. TOO RISKY!!• MboI library and FPC

Tomato Sequencing, Madison July 2006

Construction of a training set for gene prediction programa geneid

• A parameter file constructed from 112 FL sequences from different Solanaceae species (50% tomato).

• To be refined with the training set from newly released 320 FL from Shibata’s lab

Tomato Sequencing, Madison July 2006

Geneid traioned with this parameter file applied to 6 BACs in Chr9

Tomato Sequencing, Madison July 2006

Geneid vs Geneseqer

• Geneid predicted 22 genes in the 114,526 pb C09HBa0109D11.1 BAC

• Most of the predictions are supported by ESTs results as shown by geneseqer

• Geneseqer is another gene identification tool based on the “spliced alignment” of ESTs to the genomic sequence contained in the BAC

Tomato Sequencing, Madison July 2006

Geneid with a parameter file obtained from 100 Sol sequences has been applied to 6

BACs from Chr9

Vicky FernandezSheila ZunigaAngela Perez

Francisco CamaraRoderic Guigó

Miguel A BotellaAntonio Granell

Tomato Sequencing, SOL Madison, July 2006

European Commission EU-SOL