Developing a Genotyping by Sequencing Protocol for Linkage Map Construction in Black Raspberry

Since the early 1900s, the black raspberry (Rubus occidentalis L.) industry in the United States has steadily declined due to lack of adapted and disease resistant cultivars. Renewed interest in production and breeding new cultivars has been fueled by research into the use of black raspberry bioactive compounds as potential chemopreventive agents for certain cancers. We are building the genomic infrastructure for black raspberry by developing, and making available, genomic tools including molecular markers for construction of linkage and physical maps, and a draft genome assembly that will benefit both black and red raspberry U.S. breeding programs. A genotyping by sequencing (GBS) library was constructed for 92 progeny of one mapping population. The library fragment sizes ranged from 191–551 base pairs (bp) with enrichment for fragments of 191–276 bp. Single-end sequencing of 101 cycles of the 96-plex library on a single flowcell channel was performed on a Hi-Seq 2000 platform. Initial variant calling analysis through a custom data pipeline identified over 23,000 SNP/indel loci. Preliminary results indicate that GBS is an appropriate approach for SNP detection in this highly-homozygous species. Validation of these SNP followed by genetic linkage mapping coupled with anchored SSR loci will be used to improve the assembly of the draft genome which is currently at 300Mbp and 2,226 scaffolds. The construction of a densely populated genetic linkage map will be used for QTL mapping of economically important traits and for comparative genomic studies with other members of Rosaceae.