Construction Of Tobacco High Density Linkage Map By RAD (Restriction Site Associated DNA) Technique

Tobacco(Nicotiana tabacum L.) is an important model system, which has been widely used in plant physiological studies and particularly useful in bioreactor. Despite its importance, only limited molecular marker resources are available for genome analysis, genetic mapping and breeding. Restriction-site associated DNA (RAD) sequencing is a powerful new method for targeted sequencing across the genomes of many individuals. This approach has broad potential for genetic analysis through linkage mapping. We constructed a RAD library using genomic DNA from a BC1 backcross population. Sequencing 196 individuals was performed on an Illumina HiSeq 2500. Two linkage maps were constructed, one with a reference genome and another one which is termed as de novo identification of SNP by RAD-seq (DISR) without a reference genome.4,138 and 2,162 SNP markers with a total length of 1,944.74 cM and 2,000.9 cM were mapped to 24 linkage groups (LGs) in the genetic maps based on reference genome and without reference respectively.Using two distinguish SNP discovery methods based on next generation RAD sequencing technology, we have respectively mapped 2,162 and 4,318 SNPs in our backcross population. This study gives an excellent example for high density linkage map construction, whether or not it has an available genome sequence, and provides saturated information for downstream genetic investigations such as quantitative trait locus (QTL) analyses, genomic selection (e.g. bioreactor suitable cultivars).