Comparative clinical sample preparation of DNA and RNA viral nucleic acids for a commercial deep sequencing system (Illumina MiSeq (R))

dc.contributorUniversidade Estadual Paulista (UNESP)
dc.creatorUllmann, Leila Sabrina
dc.creatorTozato, Claudia de Camargo
dc.creatorMalossi, Camila Dantas
dc.creatorCruz, Tais Fukuta da
dc.creatorCavalcante, Raissa Vasconcelos
dc.creatorKurissio, Jacqueline Kazue
dc.creatorCagnini, Didier Quevedo
dc.creatorRodrigues, Marianna Vaz
dc.creatorBiondo, Alexander Welker
dc.creatorAraujo, Joao Pessoa
dc.date2015-10-21T13:11:25Z
dc.date2016-10-25T21:00:00Z
dc.date2015-10-21T13:11:25Z
dc.date2016-10-25T21:00:00Z
dc.date2015-08-01
dc.date.accessioned2017-04-06T08:59:42Z
dc.date.available2017-04-06T08:59:42Z
dc.identifierJournal Of Virological Methods, v. 220, p. 60-63, 2015.
dc.identifier0166-0934
dc.identifierhttp://hdl.handle.net/11449/128604
dc.identifierhttp://acervodigital.unesp.br/handle/11449/128604
dc.identifierhttp://dx.doi.org/10.1016/j.jviromet.2015.04.009
dc.identifierWOS:000356112300012
dc.identifierhttp://www.sciencedirect.com/science/article/pii/S0166093415001457
dc.identifier.urihttp://repositorioslatinoamericanos.uchile.cl/handle/2250/939160
dc.descriptionSequence-independent methods for viral discovery have been widely used for whole genome sequencing of viruses. Different protocols for viral enrichment, library preparation and sequencing have increasingly been more available and at lower costs. However, no study to date has focused on optimization of viral sample preparation for commercial deep sequencing. Accordingly, the aim of the present study was to evaluate an In-House enzymatic protocol for double-stranded DNA (dsDNA) synthesis and also compare the use of a commercially available kit protocol (Nextera XT, Illumina Inc, San Diego, CA, USA) and its combination with a library quantitation kit (Kapa, Kapa Biosystems, Wilmington, MA, USA) for deep sequencing (Illumina Miseq). Two RNA viruses (canine distemper virus and dengue virus) and one ssDNA virus (porcine circovirus type 2) were tested with the optimized protocols. The tested method for dsDNA synthesis has shown satisfactory results and may be used in laboratory setting, particularly when enzymes are already available. Library preparation combining commercial kits (Nextera XT and Kapa) has yielded more reads and genome coverage, probably due to a lack of small fragment recovering at the normalization step of Nextera XT. In addition, libraries may be diluted or concentrated to provide increase on genome coverage with Kapa quantitation. (C) 2015 Elsevier B.V. All rights reserved.
dc.descriptionFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
dc.languageeng
dc.publisherElsevier B.V.
dc.relationJournal Of Virological Methods
dc.rightsinfo:eu-repo/semantics/openAccess
dc.subjectDouble-stranded DNA
dc.subjectLibrary preparation
dc.subjectDeep sequencing
dc.subjectMiSeq
dc.titleComparative clinical sample preparation of DNA and RNA viral nucleic acids for a commercial deep sequencing system (Illumina MiSeq (R))
dc.typeOtro


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