Phage Genomic DNA Isolation and Annotation

Genomic DNA of bacteriophages was prepared from high-titer (i.e., >10¹⁰ PFU/mL) stocks after DNase treatment using the Norgen Biotek Phage DNA Isolation Kit (Cat# 46800) according to the manufacturer’s guidelines and sequenced at the Microbial Genome Sequencing Center (MiGS, Pittsburgh, PA, USA). Sequencing libraries were prepared using the Illumina DNA Prep kit and IDT 10 bp UDI indices and sequenced on an Illumina NextSeq 2000, producing 150 bp paired-end reads. Demultiplexing, quality control, and adapter trimming were performed with bcl-convert (v3.9.3). Reads were trimmed to Q28 using BBDuk from BBTools. Phage genomes were then assembled de novo using SPAdes 3.15.2 in --careful mode with an average read coverage of 10–50×. Assembled genomes were then annotated using Prokka version 1.14.6^{52 (link)} with default parameters, except that the PHROGs HMM database^{53 (link)} was used as input to improve phage functional gene annotations.