GSM9068147: Sample11-Stool-P1; Hepatitis E virus; OTHER
Total RNA was extracted from plasma, stool suspension or urine using the Cobas AmpliPrep Total Nucleic Acid Isolation Kit (Roche, Basel, Switzerland). Complementary DNA (cDNA) synthesis and amplicon generation were conducted in accordance with Todt et al. (6). Briefly, cDNA was synthesised from 2 to 8 μL of purified total RNA using the SuperScript III first-strand synthesis system (Life Technologies, Carlsbad, CA, USA). Amplicons were generated in a two-step touchdown PCR using TaKaRa Ex Taq Hot Start Version polymerase (Dalian, China). The NEBNext Ultra II FS DNA Library Preparation Kit (NEB) was used to prepare sequencing libraries according to the manufacturer's recommendations. Library quality was determined with the Agilent High Sensitivity DNA Kit (Agilent Technologies, Waldbronn, Germany) and a 2100 Bioanalyzer Instrument (Agilent Technologies). All samples were normalised and quantified using the KAPA Library Quantification Kit for Illumina (Kapa Biosystems, Wilmington, MA, USA). Sequencing was conducted on a Illumina MiSeq using a 2 × 300 base pair (bp) paired-end sequencing protocol.