The P. leucopus were specific pathogen–free, colony-bred animals of LL Stock from the Peromyscus Genetic Stock Center (PGSC) of the University of South Carolina. The rodents were euthanized by carbon dioxide overdose and intracardiac exsanguination. DNA for library construction was obtained from the liver and kidney, homogenized in liquid nitrogen, and extracted using a Qiagen Blood and Cell Culture kit with modifications suggested in (7). For Illumina libraries, DNA was Covaris-sheared to 500 bp and prepped using a Bioo Scientific NEXTFLEX Rapid DNA-seq kit with four cycles of PCR amplification. After size selection, we collected four HiSeq 2500 PE100 lanes for a total of 158 Gb of raw data (or roughly 54×). For PacBio libraries, we followed the study of Chakraborty et al. (7) by shearing the DNA using a 1.5-inch 24-gauge blunt-tipped needle, and the library was prepared using PacBio’s SMRTbell template protocol and Blue Pippin size selection using a 15- to 50-kb cutoff. We generated several dozen libraries in this manner and collected 124 total SMRTcells of data for a total of 104 Gb with an N50 read length of 15.7 kb (~36× coverage). Two Hi-C libraries were also constructed with six cycles of PCR amplification, and PE100 was sequenced to 20× and 8× of raw coverage. On the basis of aligning back to the final assembly, this represented 341,768× and 140,112× of span coverage, respectively.

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