Application of RADseq to The Linkage Mapping of Sugarcane to Identify Markers Linked to Early Ripening

Restriction associated DNA sequencing (RADseq) was investigated as a high-throughput SNP genotyping platform to identify markers linked to early ripening in sugarcane. This involved the reduction of the complexity of the sugarcane genome by a double digest followed by the sequencing of a subset of the restricted fragments using the Illumina Hiseq 2000. The mapping parents CP 67412 (early ripening) and M 245/76 (late) and 107 derived progenies were subject to RADseq genotyping in two sequencing libraries. The sequences were analyzed using a modified version of Stacks software (www.creskolab.uoregon.edu/stacks) using sorghum as reference genome. Nearly 9,500 SNP markers were scored for the mapping parent CP 67412 and 7,560 were scored for the late ripening variety M 245/76. Analysis of SNP marker data using Joinmap 4.0 produced linkage maps of 145 linkage groups (LGs) for variety CP 67412 and 156 LGs for variety M 245/75 with a total map length of 5, 831cM and 7,634 cM respectively. Both maps harbored more than 3,200 markers with an average marker distance of 1.7 cM and 2.1 cM respectively. LGs were thus classified into homology groups (HGs) based on their collinearity to sorghum. Even distribution of the LGs across the ten HGs representing the ten sorghum chromosomes suggested a good coverage of the sugarcane genome. High throughput, cost effectiveness and automated SNP marker discovery and scoring makes RADseq the marker of choice for sugarcane linkage map construction.