Nexteraxt guide

Genomic DNA was extracted from the 40 E. coli isolates from 2015 using an Invitrogen Easy-DNA Kit (Invitrogen, Carlsbad, CA, United States) and DNA concentration was determined using the Qubit dsDNA BR assay kit (Invitrogen). The genomic DNA was prepared for Illumina pair-end sequencing using the Illumina (Illumina, Inc., San Diego, CA) NexteraXT^® Guide 14 150319425031942 following the protocol revision C. The libraries were sequenced using an Illumina MiSeq platform. Raw sequence data were submitted to the European Nucleotide Archive (ENA) under study accession no: PRJEB30991. Along with the 2018 genomes retracted from ENA under the project accession PRJEB37980., the raw reads of all the 114 strains were de novo assembled using SPAdes algorithm for de novo short reads assembly (Bankevich et al., 2012 (link)). Assembled sequences were analyzed using the CGE tools³ including MLST finder to determine E. coli multi-locus sequence types (MLST), ResFinder 3.1 for detection of genes and chromosomal mutations mediating AMR, VirulenceFinder 2.0 with default settings for detection of virulence genes, PlasmidFinder for detection of plasmid replicons, and pMLST for further subtyping of specific plasmids.

The E. coli EC1945 isolate genome was sequenced at EURL-AR, DTU, Lyngby, Denmark, under the scope of the EFSA confirmatory testing [36 (link)]. Genomic DNA was extracted using an Invitrogen Easy-DNA KitTM (Invitrogen, Carlsbad, CA, United States), and the DNA concentrations were determined using the Qubit dsDNA BR assay kit (Invitrogen). Genomic DNA was prepared for Illumina pair-end sequencing using the Illumina (Illumina, Inc., San Diego, CA, USA) Nextera XT^® Guide following the protocol revision C1. A sample of the pooled Nextera XT Libraries was loaded onto an Illumina MiSeq reagent cartridge using MiSeq Reagent Kit v3. The libraries were sequenced using an Illumina MiSeq platform (Illumina). The raw reads were de novo assembled using the assembler pipeline (version 1.4) available from the Center for Genomic Epidemiology (CGE) (https://www.genomicepidemiology.org/) (accessed on 25 April 2022).
The Whole Genome Shotgun project has been deposited at DDBJ/ENA/GenBank under the accession JAUTEE000000000.

V. choleare genomic DNA was extracted using the Invitrogen Easy-DNA^TM Kit (Invitrogen, Carlsbad, CA, USA). The concentrations of the extracted DNA were determined using a Qubit dsDNA BR assay kit (Invitrogen). The genomic DNA was prepared for Illumina paired-end sequencing using the Illumina (Illumina, Inc., San Diego, CA) NexteraXT® Guide 150319425031942 following protocol revision C. A sample of pooled NexteraXT Libraries was loaded onto an Illumina MiSeq reagent cartridge using MiSeq Reagent Kit v2 and 500 cycles with a Standard Flow Cell. The libraries were sequenced using the MiSeq Illumina platform and MiSeq Control Software 2.3.0.3. All strains were paired-end sequenced.
Raw sequence data were submitted to the European Nucleotide Archive (http://www.ebi.ac.uk/ena) under study accession no.: PRJEB14630 (http://www.ebi.ac.uk/ena/data/view/PRJEB14630). The raw reads were assembled using the Assemble pipeline (version 1.0) available from the Center for Genomic Epidemiology (CGE; http://cge.cbs.dtu.dk/services/all.php) based on the Velvet algorithms for de novo short reads assembly. A complete list of genomic sequence data is available in Table B in S1 File.