Hordeum vulgare subsp. vulgare cultivar:CI 16139 Genome sequencing and assembly

Our goal is to resolve the Reaction to Puccinia striiformis8 (Rps8) locus on chromosome 4H in the barley accession CI 16139. Chromosome flow sorting of CI 16139 chromosome 4H was performed using the methods described by Dolezel et al. (2012) Functional and Integrative Genomics 12, 397-416 and chromosomal high molecular weight (HMW) DNA was prepared as described in Thind et al. (2017) Nature Biotechnology 35, 793-796. Chicago Dovetail sequencing of the chromosome was performed by Dovetail Genomics (Santa Cruz, CA, USA), with initial assembly in Meraculous and final scaffolding in HiRise. Briefly, Chicago libraries were generated by in vitro chromatin reconstitution using 250 ng HMW DNA. After fixation with formaldehyde, chromatin was digested with MboI, biotinylated nucleotides were added to 5 prime overhangs, and proximity ligation was performed. After reversal of crosslinking the DNA was treated to remove biotin and sheared to approximately 400 bp. A sequencing library was generated using NEBNext Ultra enzymes (New England Biolabs) and Illumina-compatible adapters and sequenced using Illumina HighSeq X to produce 229 million paired end reads, which gave 95x coverage for chromosome 4H. An initial assembly using Meraculous had length 521.7 Mb on 57,043 scaffolds. The HiRise assembly had 527.17 Mb on 1,702 scaffolds.