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Www.sistemasgenomicos.com 2nd TOMATO FINISHING WORKSHOP chromosome 9 Wageningen, April 24-25, 2008.

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Presentation on theme: "Www.sistemasgenomicos.com 2nd TOMATO FINISHING WORKSHOP chromosome 9 Wageningen, April 24-25, 2008."— Presentation transcript:

1 www.sistemasgenomicos.com 2nd TOMATO FINISHING WORKSHOP chromosome 9 Wageningen, April 24-25, 2008

2 www.sistemasgenomicos.com Originally, 142 MARKERS -Most of them in heterocromatin -Very few associated to BACs -Gap of 46cM New BACs anchored to chr9: -Syngenta data -Filter hybridization -IL mapping of BACs at ramdom CURRENT STATUS OF CHROMOSOME 9

3 www.sistemasgenomicos.com COMPLETE 65 BACs IN PROGRESS 9 BACs VERIFYING 35 BACs CURRENT STATUS OF CHROMOSOME 9 2 BACs sequencing 7 BACs in finishing pipeline 13 phosmids 12 FPC data 1 seed 5 mapping IL (chr12) 4 extending BACs 12 BACs en HTG 1 33 BACs en HTG 2 20 BACs en HTG 3 34% done

4 www.sistemasgenomicos.com SEQUENCING AND FINISHING PIPELINE BAC 7098781nt sequenced 6040781nt non-redundant ~109 kb/BAC ~15 kb overlap/BAC validation subcloning puC19 extraction, fragmentation & reparation puC19 sequencing (Big-dye terminator, AB3730) contigs Phred, Phrap, Consed PCR or direct sequencing read pair info electronic digestion whole sequence

5 www.sistemasgenomicos.com IDENTIFYING NEW BACs BLASTN against BAC end database IF no candidates found OR overlaps too large THEN oligo design at BAC ends in order to hybridize filters FILTER HYBRIDIZATION with markers not associated to BACs IL MAPPING of BACs at random (200 BACs/chr)?? FPC data from HindIII and MboI libraries at Sanger Institute Not very useful: Till now, either our BACs were not in a FPC contig or FPC contigs were too poor to find candidates

6 www.sistemasgenomicos.com GS-FLX: 11 BACs were sequenced with 454-Roche technology Elegibility criteria: size, difficulty (repeats, poli-A/T tails…), HTG phase Results (100kb BAC, 15-20x): Goods -No risk of loosing parts during library construction -In general, gaps are small -It resolves A-T poli-tails but only if the coverage is high. Bads -If repeats are present the number of contigs increases substantially -Sequencing stops when AT is repeated several times -If ex. AAATTTTTTTTAAAAA, bases at both sides are not clear unless coverage is high NEW SEQUENCING TECHNOLOGIES

7 www.sistemasgenomicos.com Some tomato jokes: There were three tomatoes: a mum, a dad and a son. The son lagged behind and fell splat on the floor. His dad yelled to him, “Ketchup, son”. A faucet, lettuce and a tomato were in a race... what happened? The faucet was running, the lettuce was ahead, and the tomato was trying to ketchup! What is a tomato with a cloak? A super-tomato What is a potato with a cloak? A potato with a super-tomato costume INSTEAD OF A RECIPE…

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