About: A Novel Method for the Capture-based Purification of Whole Viral Native RNA Genomes

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An Entity of Type : schema:ScholarlyArticle, within Data Space : covidontheweb.inria.fr associated with source document(s)

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type	Academic Article research paper schema:ScholarlyArticle
isDefinedBy	Covid-on-the-Web dataset
title	A Novel Method for the Capture-based Purification of Whole Viral Native RNA Genomes
Creator	Maurer-Stroh, Sebastian Wan, Yue Sessions, October Tan, Cedric Florez De Sessions, Paola »more»
source	BioRxiv; MedRxiv
abstract	Current technologies for targeted characterization and manipulation of viral RNA either involve amplification or ultracentrifugation with isopycnic gradients of viral particles to decrease host RNA background. The former strategy is non-compatible for characterizing properties innate to RNA strands such as secondary structure, RNA-RNA interactions, and also for nanopore direct RNA sequencing involving the sequencing of native RNA strands. The latter strategy, ultracentrifugation, causes loss in genomic information due to its inability to retrieve unassembled viral RNA. We developed a novel nucleic acid manipulation technique involving the capture of whole viral native RNA genomes for downstream RNA assays using hybridization baits in solution to circumvent these problems. This technique involves hybridization of biotinylated baits at 500 nucleotides (nt) intervals, stringent washes and release of free native RNA strands using DNase I treatment, with a turnaround time of about 6 h 15 min. Proof of concept was primarily done using RT-qPCR with dengue virus infected Huh-7 cells. We report that this protocol was able to purify viral RNA (561-791 fold). We also describe a successful application of our capture-based purification method to direct RNA sequencing, with a 77.47% of reads mapping to the target viral genome. We observed a reduction in human host RNA background by 1580 fold, a 99.91% recovery of viral genome with at least 15x coverage, and a mean coverage across the genome of 120x. This report is, to the best of our knowledge, the first description of a capture-based purification method for whole viral RNA genomes. The fundamental advantages of using our capture-based purification method makes it a superior alternative to conventional viral purification methods and would potentially pave a new path for the direct characterization and sequencing of native RNA molecules.
has issue date	2018-09-22(xsd:dateTime)
bibo:doi	10.1101/410282
has license	biorxiv
sha1sum (hex)	4c6f59325d77d7b1fc4263fc6992589a8d88db9d
schema:url	https://doi.org/10.1101/410282
resource representing a document's title	A Novel Method for the Capture-based Purification of Whole Viral Native RNA Genomes
schema:publication	bioRxiv
resource representing a document's body	covid:4c6f59325d77d7b1fc4263fc6992589a8d88db9d#body_text
is schema:about of	named entity 'strands' named entity 'gradients' named entity 'RNA' named entity 'SEQUENCING' named entity 'NON-' »more»

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