RNA-Seq for homo sapiens: serum
Total RNA was purified from 200 L of serum samples, using the PureLink Viral RNA/DNA Mini Kit (Invitrogen, USA), following the manufacturers recommendations. The complementary DNA (cDNA) synthesized for the RNA was amplified by a random PCR as described previously (Khan et al., 2016). This amplification step had the aim of increasing the viral genome for nucleotide sequencing. The PCR products were purified using the QIAquick PCR Purification Kit (QIAGEN, USA), following the manufacturers recommendations. DNA concentration was determined with the NanoDrop 2000 (Thermo Scientific, USA). PCR products of a serum sample or PCR pools containing 2 to 4 samples each, in equimolar concentration, were used for nucleotide sequencing. The sequencing libraries were prepared using the Nextera XT DNA Library Preparation Kit (Illumina, San Diego, USA), according to the manufacturers recommendations. The samples were labeled with adapter sequences using the Nextera XT Index Kit (Illumina, San Diego, USA), according to the manufacturers recommendations. Paired-end sequencing was performed using the Illumina MiSeq platform (Illumina, USA) and the MiniSeq Reagent Nano kit V2 (300-cycles) (Illumina, USA).