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CHROMOSOME 12

CHROMOSOME 12. Giovanni Giuliano PAG, 2006. Tomato chr. 12. Principal Investigator s Luigi Frusciante (UniNa) Giovanni Giuliano (ENEA) Giorgio Valle (CRIBI) Other Investigator s Amalia Barone ( UniNa) Francesca Cecchi (BIOGEN) Maria Luisa Chiusano (UniNa) Mara Ercolano (UniNa)

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CHROMOSOME 12

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  1. CHROMOSOME 12 Giovanni Giuliano PAG, 2006

  2. Tomato chr. 12 Principal Investigators Luigi Frusciante (UniNa) Giovanni Giuliano (ENEA) Giorgio Valle (CRIBI) Other Investigators AmaliaBarone (UniNa) Francesca Cecchi (BIOGEN) Maria Luisa Chiusano(UniNa) MaraErcolano (UniNa) Silvana Grandillo(IGV-CNR) Renato Rodrigues-Pousada (ENEA) Alessandro Vezzi (UniPd) Funding Agronanotech (Italian Ministry of Agriculture - started October, 2004) FIRB (Italian Ministry of Research - started October, 2005) EU-SOL (European Commission - starting April 2006)

  3. SEQUENCING CRIBI Enea BAC repository Unina Sequence assembly and Annotation Data repository

  4. Validation at UniNA • -Single colony picking • -Medium scale prep • Restriction analysis to check E. coli contamination • BAC-end sequencing -PCR markers development • Amplicon sequencing and sequence alignment • -Mapping on ILs -FISH on selected BACS

  5. Marker Bac analyzed Bac end sequencing Cornell matching Specific amplicon Amplicon Sequence match IL mapping Fish mapping Sequencing status t1211 LeHba021L02 in pipeline t989 LeHba032k07 LeHba015A04 - finished cLPT-E9 LeHba026c1 Pd LeHba026c13v LeHba026c13n (1) (1) in pipeline C2At4g03280 LeHba140m01 - in pipeline t1045 LeHba161H10 in pipeline t1487 LeHba260c13 - in pipeline t1481 LeHba075c18 in pipeline t1667 LeHba146I19 LeHba244C09 (1) (1) - in pipeline clet-8k4 LeHba189E07 LeHba161b22 Poor - tg 68 LeHba152M18 Difficult tg 263 LeHba244J04 - Difficult t0028 LeHba163O04 in pipeline ssr124 LeHba059A05 LeHba061j21 Results (1) (1) (1) finished yes One BAC end (1) may be ? Contradictory data no

  6. Validation at ENEA • -Single colony picking • -Medium scale prep (problems w minipreps) • Digestion with restriction enzymes to check E. coli contamination • End sequencing + sequencing with 2 internal oligos • (passes test if 3 sequences out of 4 are good and matching)

  7. Results Confirmation Validation by internal sequencing marker End sequence offset clone name Cornell end seqs? T0770 101 LE_HBa0183M06 Y(T7/SP6) Y(both) Yes (primer 5' to overgo ) T0770 101 LE_HBa0155J24 Y(T7/SP6) Y(T7) Yes (primer 5' to overgo ) T1676 86 LE_HBa0115G22 Y(T7/SP6) Y(both) Yes (primer 3' to overgo ) T1266 71 LE_HBa0193C03 Y(T7/SP6) Y(both) Yes (primer 5' to overgo ) TG350 120 LE_HBa0053L10 Y(T7/SP6) Y(both) To be rechecked T1504 115 LE_HBa0147G13 Y(T7/SP6) Y(both) To be rechecked T1266 71 LE_HBa0079O22 Y(T7) Y(T7) No TG394 68 LE_HBa0107D15 Y(T7/SP6) not possible No SSR124 60 LE_HBa0061J21 Y(T7/SP6) Y(both) No P62 57,2 LE_HBa0104O23 Y(T7/SP6) Y(both) No 3 BAC’s validated (3 markers); 5 BAC’s not validated (4 markers); 2 BAC’s to be rechecked (2 markers)

  8. LeHba140m01 LeHba026c13v LeHba260c13 8 in pipeline LeHba075c18 LeHba163O04 LeHba032k07 LeHba244C09 LeHba161H10 LeHba021L02 2 finished LeHba059A05 LE_HBa0193C03 LE_HBa0115G22 3 validated LE_HBa0183M06 Summary

  9. Sequencing and assembly (CRIBI)

  10. Bioinformatics at UniNA • Annotated EST database from dbEST (NCBI)* for Tomato and Potato species (updated December 2005) • Gbrowse interface for BAC annotation ready. To be released to the SOL community. * D'AGOSTINO, AVERSANO, CHIUSANO ML,, “ParPEST: A PARALLEL PIPELINE FOR EST DATA ANALYSIS ”; BMC Bioinformatics 1 December 2005.

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