Assignment of Draft Sequences of the Radish (Raphanus sativus L.) Genome to a High-Density Linkage Map

Radish (Raphanus sativus L., n=9) is one of the major vegetables in Asia.  Using an Aokubi-inbred line (‘Aokubi’), which is Japanese radish with a long and thick root, genome sequencing was performed by Next Generation Sequencer (NGS). The short reads of genomic sequences of 191.1 Gb were de novo assembled along with the BAC-end sequences. Totally, 76,592 scaffolds of more than 300 bp were constructed. The whole draft genomic sequence was 402 Mb (N50: 46,262 bases) spanning 75.9% of the estimated genomic size and contained 61,572 predicted genes. To construct a radish linkage map, we developed 746 EST-SNP markers using an F₂ population derived from cross between ‘Aokubi’ and an Sayatori-inbred line (‘Sayatori’) that is a seedpod vegetable with a very thin and small root. Subsequently, 221 SNP markers and 768 PCR-RFLP markers were produced by a comparison of SNPs between ‘Sayatori’ reads sequenced by NGS and ‘Aokubi’-scaffold sequences, and were also used for construction of a radish map.  The map was combined further with another radish linkage map constructed mainly with EST-SSR markers into a high-density integrated map of 1,166 cM with 2,553 DNA markers. The frequency of mis-assembly of scaffolds was estimated to be ca. 5% and 1,345 of the 76,592 scaffolds were assigned to the integrated map using alignments with the marker sequences, spanning 116.0Mb.  To assign more scaffolds with more than 50 kb, genotyping by NGS using primers designed from sequences of the scaffolds is in progress for validation of assembly of the scaffolds.