Genomic DNA was sequenced by SNPsaurus (Eugene, OR) on an Illumina HiSeq 4000 (Illumina, San Diego, CA), following their standard workflow for library preparation and read trimming. A Nextera tagmentation kit (Illumina) was used for library generation, followed by paired-end 150-bp sequencing at greater than 60× coverage for each of the eight isolates. Adaptors were trimmed from the reads with BBDuk (50 (link)).
To obtain long reads for scaffolding, we also sequenced the DNA on an Oxford Nanopore MinION (Oxford Nanopore, England, UK). We prepared 1 μg of DNA per isolate using the rapid sequencing kit (RAD004; Oxford Nanopore), and all DNA was unsheared prior to library preparation. We ran four isolates per flow cell sequentially. Although the flow cell was washed after each isolate, we barcoded each with a rapid barcoding kit (RBK004; Oxford Nanopore) to guard against cross-contamination. Reads were called during sequencing using Guppy v2.0.10, and only the reads which passed the initial quality check during the run (and which were therefore placed into the “fast5_pass” folder) were used (51 (link)).
To obtain long reads for scaffolding, we also sequenced the DNA on an Oxford Nanopore MinION (Oxford Nanopore, England, UK). We prepared 1 μg of DNA per isolate using the rapid sequencing kit (RAD004; Oxford Nanopore), and all DNA was unsheared prior to library preparation. We ran four isolates per flow cell sequentially. Although the flow cell was washed after each isolate, we barcoded each with a rapid barcoding kit (RBK004; Oxford Nanopore) to guard against cross-contamination. Reads were called during sequencing using Guppy v2.0.10, and only the reads which passed the initial quality check during the run (and which were therefore placed into the “fast5_pass” folder) were used (51 (link)).