Sequencing update of tomato chromosome 3 Chinese Academy of Sciences Chuanyou Li Chinese Academy of Sciences 2019/1/18

Physical map construction FISH BACs sequencing Problems

BAC libraries Three BAC libraries of tomato were ordered and duplicated. (HindIII, EcoRI, MboI) Manual editing of the available FPC data: 7000 contigs---3000 contigs

Physical map construction critical for whole genome sequencing ~1000 genetic markers don't have corresponding BACs physical map coverage is very low only ~50Mb of 950 Mb of tomato genome physical map is not focused on the euchromatin region no minimum tilling path could be developed at this moment sequencing has to step-by-step, BAC-by-BAC

PCR screening of Positive BACs Containing the 986 Genetic Markers PCR of Heinz PCR of superpool PCR of each pool Data deconvolution individual BAC confirmation pool plate 1 pool plate 2 pool plate 3 pool plate 4 pool plate 5

Screening and confirmation of markers Chr. makers Tested confirmed 1 133 102 68 2 119 83 53 3 81 73 49 4 110 90 57 5 67 47 21 6 75 39 7 92 52 8 69 54 28 9 76 55 35 10 62 50 45 11 56 22 12 33 sum 985 756 480

Library quality Too many hits No hits This effort provides little help to refine the physical map.

Tomato sequence FISH Institute of Genetics and Developmental Biology, Chinese Academy of Sciences

The borderlines on the short and long arm of tomato chromosome 3 The order of clones on tomato chromosome 3 Heterochromatin and euchromatin study on tomato chromosome 3

Chinese Group Target Tomato chromosome 3 Short arm cen Long arm 348--47--20

The clone of TG479 located at the end of short arm and the clone of T1540 on borderline between the short arm heterochromatin and euchromatin.

The clone of TG244 located at the end of long arm

The nearest clone of TG247 located on the long arm heterochromatin from centromere.

The clone of T0772 located on boundary between the heterochromatin and euchromatin of the long arm.

The order and distance of clones on tomato chr3 T1235 160cM SSR11 164cM T0761 133cM T0482 137cM

The order and distance of clones on tomato chr3 T1511 92cM TG284 92.3 or 123.0cM

Based on FISH analysis, 47 seed BACs were confirmed.

BACs sequencing 3730 standard, 10-15X coverage 97 BACs were on the sequencing pipeline 44 BACs were finished. 15 BACs were submitted to SGN.

Problems: Physical map? Identification of more BACs?

Acknowledgment SOL project committee The Ministry of Science and Technology of China. Institute of Genetics and Development Biology, CAS Wuhan Botanical Garden, CAS National Center for Gene Research, CAS Shandong Agricultural University, China Mingsheng Chen, Zhukuan Cheng, Hongqing Ling, Jianru Zuo, Yongbiao Xue,Ying Wang, Bin Han

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