Nexseq 500 system

Total EV-associated miRNA of 24 samples in the derivation set was isolated using the miRNeasy Mini Kit (QIAGEN, The Netherlands) and sent to DNA Chip Research Inc., where small RNA library preparation and sequencing were performed. Small RNA libraries were prepared using the QIAseq miRNA library kit and QIAseq miRNA NGS 96 Index IL for Illumina, and yields were evaluated using the Agilent 2100 BioAnalyzer. Small RNA sequencing was performed using the Illumina NexSeq 500 system.

Fusion genes in recombinant vectors were verified by next-generation sequencing on a NexSeq 500 system (Illumina, San Diego, CA, USA). Specific sequencing primers were obtained from the aLICator LIC Cloning and Expression Kit, and the gene sequences were analyzed by BLAST and submitted to GenBank at NCBI.

The variable regions of class 1 and 2 integrons from the MDR-UPEC clinical strains were amplified by PCR using a high-fidelity Pfu polymerase of Thermo-Fisher Scientific (CA, USA) (Table S1). The amplicons were cleaned and concentrated using the Zymo DNA Clean and Concentrator of Zymo Research (CA, USA) and subjected to next-generation sequencing on a NexSeq500 System (Illumina, CA, USA), which was performed at “Unidad de Secuenciación del Instituto Nacional de Medicina Genómica” (CDMX, Mexico). The sequences were analyzed and assembled using ClustalO, ORF Finder (Open Reading Frame Finder), and BLAST (Basic Local Alignment Search Tool) from the NCBI (National Center of Biotechnology Information) (Sievers et al., 2011 (link); Soleimani et al., 2014 (link)).