Comparing viral metagenomics methods using a highly multiplexed human viral pathogens reagent.

摘要: Unbiased metagenomic sequencing holds significant potential as a diagnostic tool for the simultaneous detection of any previously genetically described viral nucleic acids in clinical samples. Viral genome sequences can also inform on likely phenotypes including drug susceptibility or neutralization serotypes. In this study, different variables laboratory methods often used to generate metagenomics libraries were compared their abilities detect multiple viruses and full coverage. A biological reagent consisting 25 human RNA DNA pathogens was estimate effect filtration nuclease digestion, DNA/RNA extraction methods, pre-amplification use library preparation kits acids. Filtration treatment led slight decreases percentage sequence reads number detected. For acid extractions silica spin columns improved recovery relative magnetic beads Trizol extraction. Pre-amplification using random RT-PCR while generating more resulted fewer viruses, overlapping sequences, lower The ScriptSeq method retrieved greater fraction genomes than TruSeq Nextera methods. able simultaneously up 22 analyzed all those detected by qPCR. Further optimization will be required biologically complex samples such tissues, blood, feces.