A one step real time PCR method for the quantification of hepatitis delta virus RNA using an external armored RNA standard and intrinsic internal control

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dc.contributor.authorKaratayli, Ersin
dc.contributor.authorAltunoglu, Yasemin Celik
dc.contributor.authorKaratayli, Senem Ceren
dc.contributor.authorAlagoz, S. Gokce K.
dc.contributor.authorCinar, Kubilay
dc.contributor.authorYalcin, Kendal
dc.contributor.authorIdilman, Ramazan
dc.date.accessioned2024-04-24T16:15:02Z
dc.date.available2024-04-24T16:15:02Z
dc.date.issued2014
dc.departmentDicle Üniversitesien_US
dc.description.abstractBackground: Hepatitis delta virus (HDV) RNA viral load measurement is critical in diagnosis and monitoring the response to antiviral treatment. Objectives: Our aim is to design a real time PCR method for accurate quantitation of HDV RNA in clinical specimens using an armored RNA as external standard, and an intrinsic internal control. Study design: A plasmid bearing delta antigen region of genotype I HDV genome was used to develop an armored RNA. Serial dilutions of the armored HDV RNA standard with 10(12) copy/mL were used as standards for quantitation. A primer-probe set derived from HDAg region was used in one step EZ RT PCR kit chemistry which uses rTth enzyme allowing reverse transcription and polymerization in the same tube. The kit also uses the advantage of uracil-N-glycosylase (UNG) enzyme treatment to prevent PCR contamination. Results: The established assay has a dynamic range of 10(2)-10(11) copy/mL with a PCR efficiency of 96.9%. Detection limit was 858 +/- 32 copy/mL with 95% confidence interval. Intra-and inter-assay variabilities were low for high, medium and low levels of viremia. Incorporation of freely circulating GAPDH in serum into the assay as an intrinsic internal control prevented false negative results and failures in PCR amplifications due to inhibitors, inefficient extraction procedures or enzymatic reactions. Conclusion: In conclusion, this study defines a novel assay for sensitive and reliable quantification of HDV RNA using an armored HDV RNA as a standard and GAPDH in plasma or serum as an intrinsic internal control in a single tube. (c) 2014 Elsevier B.V. All rights reserved.en_US
dc.description.sponsorshipAnkara University Scientific Research Projects [11B3330005]en_US
dc.description.sponsorshipThis study is supported by Ankara University Scientific Research Projects (project no: 11B3330005).en_US
dc.identifier.doi10.1016/j.jcv.2014.01.021
dc.identifier.endpage15en_US
dc.identifier.issn1386-6532
dc.identifier.issn1873-5967
dc.identifier.issue1en_US
dc.identifier.pmid24594080
dc.identifier.scopus2-s2.0-84899077454
dc.identifier.scopusqualityQ1
dc.identifier.startpage11en_US
dc.identifier.urihttps://doi.org/10.1016/j.jcv.2014.01.021
dc.identifier.urihttps://hdl.handle.net/11468/15611
dc.identifier.volume60en_US
dc.identifier.wosWOS:000334510800003
dc.identifier.wosqualityQ2
dc.indekslendigikaynakWeb of Science
dc.indekslendigikaynakScopus
dc.indekslendigikaynakPubMed
dc.language.isoenen_US
dc.publisherElsevier Science Bven_US
dc.relation.ispartofJournal of Clinical Virology
dc.relation.publicationcategoryMakale - Uluslararası Hakemli Dergi - Kurum Öğretim Elemanıen_US
dc.rightsinfo:eu-repo/semantics/closedAccessen_US
dc.subjectReal Time Pcren_US
dc.subjectHdven_US
dc.subjectArmored Rnaen_US
dc.subjectIntrinsic Controlen_US
dc.titleA one step real time PCR method for the quantification of hepatitis delta virus RNA using an external armored RNA standard and intrinsic internal controlen_US
dc.titleA one step real time PCR method for the quantification of hepatitis delta virus RNA using an external armored RNA standard and intrinsic internal control
dc.typeArticleen_US

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