Bluepippin system

To obtain a representative, high-quality reference genome for short-read mapping, S. Concord isolate ITM_8091960 was sequenced using the PacBio RSII system (Pacific Biosciences, California, USA). This isolate originated from a stool sample of an Ethiopian child that presented at the travel clinic of ITM Antwerp in 2008. DNA was prepared using the PacBio Template Prep Kit (Pacific Biosciences, California, USA) and the BluePippin™ system for size selection, for sequencing with the PacBio RSII system at the Wellcome Sanger Institute (Hinxton, UK). Reads were assembled de novo using the HGAP^{56 (link)} protocol v3.0 implemented in smrtanalysis v2.3.0, resulting in one chromosome-sized contig, and five small contigs. Circlator^{57 (link)} v1.5.5 was used to remove self-compatible ends and rearrange the start positions of the contigs at the dnaA gene, or a predicted gene. The resulting contigs were polished twice using Quiver^{56 (link)}.

Genomic DNA from P. savastanoi strain MHT1 was extracted with the TIANamp Bacteria DNA Kit (TIANGEN BIOTECH, Beijing, China) and quality-tested on a Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA). DNA concentration was measured on a Nanodrop (Thermo Fisher Scientific, Wilmington, USA).
Genome sequencing was performed on a PacBio long-read sequencer (Pacific Biosciences, Menlo Park, CA, USA). SMRTbell libraries were obtained using g-TUBE (Covaris, Woburn, Massachusetts, USA) and end repair. According to the manufacturer’s protocols of the Blue Pippin system (Pacific Biosciences), fragments with sizes larger than 10 kb were selected. The quality and average size of the fragments of the library were estimated using a Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA) and Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, USA). The PacBio Sequel system (Pacific Biosciences, Menlo Park, CA, USA) was used to perform SMRT sequencing. The resulting continuous long reads were used for de novo assembly with the Falcon program (version 0.3.0) [36 (link)].