Illumina MiSeq paired end sequencing; MiSeq analysis of viral progeny at 24 and 72 hours post-infection with H/PF/2013 Zika virus of HuH-7 cells
MiSeq analysis of viral progeny at 24 and 72 hours post-infection with H/PF/2013 Zika virus of HuH-7 cells
Huh-7 cells were seeded at 1x105 cells per well in 24-well tissue culture plates one day prior to infection. Cells were then infected with H/PF/2013 at a multiplicity of infection (MOI) of 1.0 by adsorbing the viral inoculum for an hour at 37°C. Inoculum was then removed and replaced with DMEM maintenance media (2% FCS, 100U/mL penicillin and 100 μg/mL streptomycin). At 24 and 72hpi, the infected cells were harvested.  Viral RNA was extracted from the supernatant using the QIAamp Viral RNA Mini kit (Qiagen) without the use of carrier RNA as per manufacturer's instructions.  NGS libraries were generated using the NEBNext Ultra Directional RNA Library Prep Kit for Illumina from NEB as per manufacturer's instructions.