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Sequencing of 6.7 Mb of the melon genome using a BAC pooling strategy González, Víctor M Benjak, Andrej Hénaff, Elizabeth Marie Mir Arnau, Gisela Casacuberta, Josep M Garcia-Mas, Jordi Puigdomènech, Pere Producció Vegetal Genòmica i Biotecnologia 575 633 Background: Cucumis melo (melon) belongs to the Cucurbitaceae family, whose economic importance among horticulture crops is second only to Solanaceae. Melon has a high intra-specific genetic variation, morphologic diversity and a small genome size (454 Mb), which make it suitable for a great variety of molecular and genetic studies. A number of genetic and genomic resources have already been developed, such as several genetic maps, BAC genomic libraries, a BAC-based physical map and EST collections. Sequence information would be invaluable to complete the picture of the melon genomic landscape, furthering our understanding of this species’ evolution from its relatives and providing an important genetic tool. However, to this day there is little sequence data available, only a few melon genes and genomic regions are deposited in public databases. The development of massively parallel sequencing methods allows envisaging new strategies to obtain long fragments of genomic sequence at higher speed and lower cost than previous Sanger-based methods. Results: In order to gain insight into the structure of a significant portion of the melon genome we set out to perform massive sequencing of pools of BAC clones. For this, a set of 57 BAC clones from a double haploid line was sequenced in two pools with the 454 system using both shotgun and paired-end approaches. The final assembly consists of an estimated 95% of the actual size of the melon BAC clones, with most likely complete sequences for 50 of the BACs, and a total sequence coverage of 39x. The accuracy of the assembly was assessed by comparing the previously available Sanger sequence of one of the BACs against its 454 sequence, and the polymorphisms found involved only 1.7 differences every 10,000 bp that were localized in 15 homopolymeric regions and two dinucleotide tandem repeats. Overall, the study provides approximately 6.7 Mb or 1.5% of the melon genome. The analysis of this new data has allowed us to gain further insight into characteristics of the melon genome such as gene density, average protein length, or microsatellite and transposon content. The annotation of the BAC sequences revealed a high degree of collinearity and protein sequence identity between melon and its close relative Cucumis sativus (cucumber). Transposon content analysis of the syntenic regions suggests that transposition activity after the split of both cucurbit species has been low in cucumber but very high in melon. Conclusions: The results presented here show that the strategy followed, which combines shotgun and BAC-end sequencing together with anchored marker information, is an excellent method for sequencing specific genomic regions, especially from relatively compact genomes such as that of melon. However, in agreement with other results, this map-based, BAC approach is confirmed to be an expensive way of sequencing a whole plant genome. Our results also provide a partial description of the melon genome’s structure. Namely, our analysis shows that the melon genome is highly collinear with the smaller one of cucumber, the size difference being mainly due to the expansion of intergenic regions and proliferation of transposable elements. We gratefully acknowledge Lifesequencing S. L. for technical assistance in 454-sequencing one of the DNA pools. This project was funded by the Plan Nacional de Investigación Científica of the Spanish Ministerio de Educación y Ciencia (Projects BIO2007-61789 to PPR and AGL2006-12780-C02-01 to JGM), by the Consolider-Ingenio 2010 Programme of the Spanish Ministerio de Ciencia e Innovación (CSD2007-00036 “Center for Research in Agrigenomics”), and by the Departament d’Innovació, Universitats i Empresa de la Generalitat de Catalunya. We acknowledge the valuable technical help from Roche 454 and Roche Spain. info:eu-repo/semantics/publishedVersion 2010-11-12 info:eu-repo/semantics/article González, Víctor M, Andrej Benjak, Elizabeth Marie Hénaff, Gisela Mir, Josep M Casacuberta, Jordi Garcia-Mas, and Pere Puigdomènech. 2010. “Sequencing of 6.7 Mb of the Melon Genome Using a BAC Pooling Strategy.” BMC Plant Biology 10 (1): 246. doi: 10.1186/1471-2229-10-246 1471-2229 http://hdl.handle.net/20.500.12327/3124 https://doi.org/10.1186/1471-2229-10-246 eng BMC Plant Biology MEC/ /BIO2007-61789/ES/Ampliación del mapa fisico y aplicación al análisis de los caracteres de forma del fruto del melón (Cucumis melo)/ MEC/ /AGL2006-12780-C02-01/ES/Caracterización agronómica y con herramientas genómicas de QTLS implicados en calidad de fruto y desarrollo de herramientas basadas en el silenciamiento génico/AGR MEC/Programa nacional de medios de transporte/CSD2007-00036/ES/Centro de Genómica Básica y de orientación Agroalimentaria/ Attribution 4.0 International http://creativecommons.org/licenses/by/4.0/ info:eu-repo/semantics/openAccess 15 BMC