Tomato Sequencing Project Meeting at SOL 2008, Oct. 15, 2008 Status of tomato genome sequence in Japan: chromosome 8 sequencing and selected BAC mixture sequencing Shusei Sato, Erika Asamizu, Takakazu Kaneko, Hiroyuki Fukuoka, Satoshi Tabata

Distribution of Anchor Markers on Chromosomes Initial seeds on Chr.8 92 165 1.8 79 143 67 171 2.6 62 137 2.2 40 119 3.0 63 101 1.6 51 112 33 87 116 2.9 41 2.1 43 103 2.4 39 120 3.1 # anchors cM chr length cM per anchor

Summary of EST-SSR Marker analysis 712 markers have been mapped on EXPEN2000 chr1 78 chr7 56 chr2 66 chr8 62 chr3 74 chr9 52 chr4 60 chr10 49 chr5 62 chr11 50 chr6 52 chr12 51 34 new seed clones have been selected

Status of Chr.8 Sequencing Nr. BACs Finished 151 Phase 2 5 Assembling 11 Sequencing 18 Total 185 (86% of initial target) (106% of initial target) Finished length without overlap: 14,906,592 bp 67 seed points (34 contigs, 11 single clones)

Gap Filling by Whole Genome Shotgun Sequencing Selected BAC Mixture (SBM) shotgun Select BACs whose end sequences do not contain undesired (repeat) sequences Mix the BACs and sequence by shotgun BAC Repeat Gene space

Both ends are NOT repeat BLASTN BES vs. RepeatDB Percentage LTR 49.7 Unknown 30.3 rRNA 7.4 Satellite 1.6 Simple repeat 1.5 DNA 1.4 LINE 1.3 402,012 end sequences from 177,408 BAC clones vs. 14,229 repeat sequences (TIGR_SolAth_repeat, mips_repeat_collection, SGN repeat collection) Library Both ends are repeat One end is repeat Both ends are NOT repeat HBa 19,123 35,277 26,181 Eco RI 9,533 18,594 15,516 Mbo I 15,570 20,995 13,134 Total 44,226 74,866 54,831 Source of the 1st BAC mixture

Selected BAC Mixture (SBM) shotgun sequencing <1st set> 10,000 clones from HBa library 5,000 clones from EcoRI library 5,000 clones from MboI library 20,000 BAC clones A total of 2.9 million sequences were accumulated by August. (total length is ~ 1.7 Gbp)

Both ends are NOT repeat BLASTN BES vs. RepeatDB Percentage LTR 49.7 Unknown 30.3 rRNA 7.4 Satellite 1.6 Simple repeat 1.5 DNA 1.4 LINE 1.3 402,012 end sequences from 177,408 BAC clones vs. 14,229 repeat sequences (TIGR_SolAth_repeat, mips_repeat_collection, SGN repeat collection) Library Both ends are repeat One end is repeat Both ends are NOT repeat HBa 19,123 35,277 26,181 Eco RI 9,533 18,594 15,516 Mbo I 15,570 20,995 13,134 Total 44,226 74,866 54,831 Source of the 2nd BAC mixture Source of the 1st BAC mixture

Selected BAC Mixture (SBM) shotgun sequencing <2nd set> 5,400 clones from HBa library 2,700 clones from EcoRI library 2,700 clones from MboI library 10,800 BAC clones Sequencing has started in September 2008.

Status of SBM Shotgun Sequences ・As of Oct. 2008, 3.0 million sequences 　has been accumulated. 　(total length is 1.9 Gbp) 3rd Assemble ・Assembled into 205,091 contigs. 　Total size of the contigs is ~ 580 Mbp. longest contig: 127,384 bp (17,702bp in 2nd Assemble) > 5kb contigs: 24,774 (21,230 in 2nd Assemble)

Status of SBM Shotgun Assembly EST coverage :Tomato Gene Index (46,849 unigenes) 74% SBM + Finished BAC (34,546 hits) 71% SBM (33,089 hits)

Blast DB of the SBM contigs > We are pleased to provide the blast DB of the SBM contigs to other groups of tomato genome sequence project for the purpose of contig extension. http://plant1.kazusa.or.jp/TBAC/blast.html user name: SOLseq password: ConExDB0804 > Please contact Shusei Sato (ssato@kazusa.or.jp) for the sequence information on the matched contigs.

Dept. Plant Genome Research National Institute of Vegetable and Tea Science Satoshi Tabata Erika Asamizu Shusei Sato Molecular Genetics and Physiology team Sequencing Marker Takakazu Kaneko Akiko Watanabe Akiko Ono Naomi Nakazaki Midori Kato Kumiko Kawashima Yoshimi Shimizu Chiharu Minami Chika Takahashi Shigemi Sasamoto Tsuyuko Wada Ai Matsuno Hiroyuki Fukuoka Satomi Negoro Yumika Kitamura