S01 - Session O5 - I - De novo assembly of Citrus limon and target-sequence genotyping towards the detection of genes involved in tolerance to ‘mal secco’ disease

Authors: Mario Di Guardo *, Marco Moretto, Mirko Moser, Chiara Catalano, Michela Troggio, Ziniu Deng, Alessandro Cestaro, Marco Caruso, Gaetano Distefano, Riccardo Russo, Silvia di Silvestro, Carmen Arlotta, Donata Pietro Paolo, Giuseppe Russo, Stefano La Malfa, Luca Bianco, Alessandra Gentile

Lemon ( Citrus limon (L.) Burm. f.) is one of the most important citrus species, mainly prized for the organoleptic and nutraceutical properties of the juice and the peel. Herein we report, for the first time, the release of a high quality reference genome of lemon. The sequencing was carried out combining Illumina and Oxford Nanopore data leading to the definition of a primary and an alternative assembly characterized by a genome size of 312.8 and 324.74 Mb, respectively. Gene annotation allowed the detection of 35,020 and 34,577 predicted transcripts, rspectively, while in silico analysis of the transposable elements (TE) allowed the identification of 2,878 and 2,897 on primary and alternative assemblies, respectively. For more than a century, 'mal secco', a tracheomycosis caused by Plenodomus tracheiphilus , has presented a major threat to lemon cultivation in many Mediterranean countries, causing the desiccation of twigs and branches until the death of the whole plant. To detect candidate genes linked to tolerance to mal secco, one inter- and two intra-specific segregating populations were obtained. Three tolerant genotypes, namely Interdonato lemon, C. clementina, and C. latipes , were crossed with the highly susceptible 'Femminello Siracusano 2Kr'. Around 300 hybrids were artificially inoculated or subjected to natural infection for two consecutive seasons, and symptoms were followed by visual screening and real-time PCR. The nown ancestors of lemon (citron, pummelo and sour orange) were also genotyped and phenotyped to achieve a more complete picture of the allelic flow through the generations. The genotyping was performed using a Single-Primer Enrichment Technology (SPET), leading to the definition of 30,000 SNPs. Two QTL approaches were carried out: a single-family QTL analysis and a pedigree-based approach (PBA) in which all accessions were analysed simultaneously, taking advantage of the availability of the pedigree records. Results shed light on the genetic regulation of the resistance to mal secco and the availability of molecular markers linked to this trait will play a crucial role to obtaining novel varieties coupling mal secco resistance with optimal fruit quality and productivity.