Chr9 A ntonio Granell IBMCP-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 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_HBa0116C Le_HBa0203J14 Le_HBa0168F Le_HBa0026P14 Le_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_EcoRI0022M12 SL_EcoRI0103M07/ SL_EcoRI0116G11 SL_EcoRI0004D19

Le_HBa0107D15 Le_HBa0099F Le_HBa0099P Le_HBa0109D11 Le_HBa0278J12 61 Le_HBa0183J Le_HBa0226D Le_HBa0165P 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_HBa0099F14 SL_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 Fernandez Sheila Zuniga Angela Perez Francisco Camara Roderic Guigó Miguel A Botella Antonio Granell Tomato Sequencing, SOL Madison, July 2006 European Commission EU-SOL