Chicago library preparation and sequencing
Dovetail Genomics LLC prepared a Chicago library as described previously (Putnam et al., 2016). Briefly, approximately 500 ng of high molecular weight gDNA (mean fragment length = 80 kb) from an individual female beetle (NCBI BioSample: SAMN14918906) was reconstituted into chromatinin vitro and fixed with formaldehyde. Fixed chromatin was digested with DpnII , the 5’-overhangs filled in with biotinylated nucleotides, and then free blunt ends were ligated. After ligation, crosslinks were reversed, and the DNA was purified from associated protein. Purified DNA was treated to remove biotin that was not internal to ligated fragments. The DNA was then sheared to approximately 350 bp mean fragment length and sequencing libraries were generated using NEBNext Ultra enzymes and Illumina-compatible adapters. Biotin-containing fragments were isolated using streptavidin beads before PCR enrichment of the library. The library was sequenced on an Illumina HiSeq X platform to produce 510 million 2x150 bp paired-end reads (NCBI SRA: SRR11965410), which provided 747 x physical coverage of the genome (1-100 kb pairs).