Building Bridges: Integrating in situ HiC and Molecular Cytogenetics to Reconstruct Rosa wichurana Pseudochromosomes

The rosa genus comprises thousands of ornamental cultivars with a great economical value (Gudin, 2000). To extend the genomic toolbox for rose breeding we initiated genome sequencing of Rosa wichurana (2n=2x=14, 0.58pg/1C, 570 Mbp, Yokoya et al., 2000), one of the species involved in the origin of many modern rose cultivars and a valuable source of resistance genes (Dugo et al. 2005; Moghaddam et al. 2012; Leus et al. 2009). 60x genome coverage of Illumina 2x250 reads was assembled using Discovar. We then used in situ Hi-C sequencing and Lachesis to create a genome contact-probability map in which 16.771 scaffolds >10kb are ordered into 7 pseudochromosomes (~0.7 genome equivalent). To anchor the draft genome structure to known linkage groups, a set of 11 reference genes have been physically positioned along the 7 pachytene chromosomes using high sensitivity Tyramide-FISH (Kirov et al., 2014; 2015), revealing good co-linearity between the cytogenetic map and the in situ Hi-C based physical map. To determine the centromere structure and position on the R. wichurana pseudochromosomes, we identified rose tandem centromeric repeat sequences in the repeatome, and positioned those by FISH on mitotic and pachytene chromosomes. Likewise, Hi-C read pairs with centromeric repeat signatures identify the centromere location in the Hi-C contact map of the pseudochromosomes. Ongoing work involves the creation of a GBS-based genetic map and further integration of complementary cytogenetic, physical and genetic maps for high-quality genome assembly of R. wichurana. The methodological aspects of the proposed pipeline and future perspectives for R. wichurana genome sequencing and annotation will be discussed.