FALCON assembly and Illumina polishing

FALCON (Chin et al. 2016) was used to generate genome assemblies for PK (v0.4.0) and FN (v1.8.6). Briefly, raw subread data were filtered to remove the shortest reads to an approximate coverage of 70× for each genome, leaving 8,003,220 (80.2%) of subreads for PK and 6,646,226 (62.6%) of subreads for FN, or ∼58 Gbp for each. Preassembled reads (i.e., error-corrected reads) were then created with a length cutoff of ≥6000 bp for PK and ≥7000 bp for FN, resulting in 2,239,051 and 5,323,023 preassembled reads, respectively. The PK and FN genomes were then assembled using preassembled reads with a minimum length of 9 kbp or 7 kbp, respectively. Additional relevant assembly parameter settings for FN were as follows:

pa_HPCdaligner_option: -B128 -t16 -e0.8 -M24 -l1200 -k18 -h256 -w8 -s100 -T12

ovlp_HPCdaligner_option: -B128 -M24 -k24 -h600 -e.92 -l1800 -s100 -T12

falcon_sense_option: --output_multi --min_cov_aln 4 --min_idt 0.70 --min_cov 4

--max_n_read 200

falcon_sense_skip_contained: False

overlap_filtering_setting: --max_diff 120 --max_cov 120 --min_cov 4

Similar assembly parameters were used for PK, except that min_cov was set to 3.

Each FALCON assembly was corrected with paired-end Illumina reads using Pilon version 1.22 (Walker et al. 2014) after mapping available Illumina sequencing data (van Bakel et al. 2011) to the FALCON-assembled genomes using BWA-MEM (version 0.7.8) (Li 2013) with an average of 96× (PK) and 23× (FN) coverage. Correction was performed with the “diploid” flag and the “bases” flag set to correct only indels and SNPs. A total of 1,511,828 insertions and 228,876 deletions were corrected in the FN assembly, and 1,807,453 insertions and 283,918 deletions were corrected in the PK assembly.