In vitro transcribed RNA molecules for the diagnosis of pandemic 2009 influenza A(H1N1) virus by real-time RT-PCR

Mario Bermúdez de León, Katia Peñuelas-Urquides, Miguel E. Aguado-Barrera, María José Currás-Tuala, Brenda L. Escobedo-Guajardo, Rosa Nelly González-Ríos, Viviana L. Mata-Tijerina, Ofelia E. Vázquez-Monsiváis

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

The 2009 influenza A(H1N1) outbreak allowed the implementation of new epidemiologic surveillance tools in several countries around the world. A new molecular protocol with appropriate sensitivity and specificity using real-time RT-PCR was developed by the Centers for Disease Control and Prevention (CDC) to identify the pandemic 2009 influenza A (H1N1) virus in human specimens. In the CDC protocol, positive controls are available only upon request and they are taken from cell cultures infected with 2009 influenza A(H1N1) virus, representing a handling risk for laboratory technicians. The poor availability of positive control materials in diagnostic laboratories may limit the public health response. The aim of the work presented in this paper was to develop positive controls for the diagnostic testing of influenza A(H1N1) virus that could be used in the CDC real-time RT-PCR protocol. A series of plasmid constructs bearing partial sequences of the viral genes were created and each construct was used as a template for in vitro transcription. RNA molecules were obtained successfully at high yield, i.e., 2×107assays per microliter. Thus, the inclusion of these molecules in the influenza panel as positive controls is proposed. The in vitro transcribed RNA could also be used as quality standards in the design of international proficiency studies. © 2013 Elsevier B.V.
Original languageEnglish
Pages (from-to)487-491
Number of pages5
JournalJournal of Virological Methods
DOIs
Publication statusPublished - 1 Nov 2013
Externally publishedYes

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H1N1 Subtype Influenza A Virus
Influenza A virus
Pandemics
Centers for Disease Control and Prevention (U.S.)
Real-Time Polymerase Chain Reaction
RNA
Human Influenza
Epidemiological Monitoring
Laboratory Personnel
Viral Genes
Disease Outbreaks
Plasmids
Public Health
Cell Culture Techniques
Sensitivity and Specificity
In Vitro Techniques

All Science Journal Classification (ASJC) codes

  • Virology

Cite this

Bermúdez de León, M., Peñuelas-Urquides, K., Aguado-Barrera, M. E., Currás-Tuala, M. J., Escobedo-Guajardo, B. L., González-Ríos, R. N., ... Vázquez-Monsiváis, O. E. (2013). In vitro transcribed RNA molecules for the diagnosis of pandemic 2009 influenza A(H1N1) virus by real-time RT-PCR. Journal of Virological Methods, 487-491. https://doi.org/10.1016/j.jviromet.2013.07.016
Bermúdez de León, Mario ; Peñuelas-Urquides, Katia ; Aguado-Barrera, Miguel E. ; Currás-Tuala, María José ; Escobedo-Guajardo, Brenda L. ; González-Ríos, Rosa Nelly ; Mata-Tijerina, Viviana L. ; Vázquez-Monsiváis, Ofelia E. / In vitro transcribed RNA molecules for the diagnosis of pandemic 2009 influenza A(H1N1) virus by real-time RT-PCR. In: Journal of Virological Methods. 2013 ; pp. 487-491.
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Bermúdez de León, M, Peñuelas-Urquides, K, Aguado-Barrera, ME, Currás-Tuala, MJ, Escobedo-Guajardo, BL, González-Ríos, RN, Mata-Tijerina, VL & Vázquez-Monsiváis, OE 2013, 'In vitro transcribed RNA molecules for the diagnosis of pandemic 2009 influenza A(H1N1) virus by real-time RT-PCR', Journal of Virological Methods, pp. 487-491. https://doi.org/10.1016/j.jviromet.2013.07.016

In vitro transcribed RNA molecules for the diagnosis of pandemic 2009 influenza A(H1N1) virus by real-time RT-PCR. / Bermúdez de León, Mario; Peñuelas-Urquides, Katia; Aguado-Barrera, Miguel E.; Currás-Tuala, María José; Escobedo-Guajardo, Brenda L.; González-Ríos, Rosa Nelly; Mata-Tijerina, Viviana L.; Vázquez-Monsiváis, Ofelia E.

In: Journal of Virological Methods, 01.11.2013, p. 487-491.

Research output: Contribution to journalArticle

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T1 - In vitro transcribed RNA molecules for the diagnosis of pandemic 2009 influenza A(H1N1) virus by real-time RT-PCR

AU - Bermúdez de León, Mario

AU - Peñuelas-Urquides, Katia

AU - Aguado-Barrera, Miguel E.

AU - Currás-Tuala, María José

AU - Escobedo-Guajardo, Brenda L.

AU - González-Ríos, Rosa Nelly

AU - Mata-Tijerina, Viviana L.

AU - Vázquez-Monsiváis, Ofelia E.

PY - 2013/11/1

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N2 - The 2009 influenza A(H1N1) outbreak allowed the implementation of new epidemiologic surveillance tools in several countries around the world. A new molecular protocol with appropriate sensitivity and specificity using real-time RT-PCR was developed by the Centers for Disease Control and Prevention (CDC) to identify the pandemic 2009 influenza A (H1N1) virus in human specimens. In the CDC protocol, positive controls are available only upon request and they are taken from cell cultures infected with 2009 influenza A(H1N1) virus, representing a handling risk for laboratory technicians. The poor availability of positive control materials in diagnostic laboratories may limit the public health response. The aim of the work presented in this paper was to develop positive controls for the diagnostic testing of influenza A(H1N1) virus that could be used in the CDC real-time RT-PCR protocol. A series of plasmid constructs bearing partial sequences of the viral genes were created and each construct was used as a template for in vitro transcription. RNA molecules were obtained successfully at high yield, i.e., 2×107assays per microliter. Thus, the inclusion of these molecules in the influenza panel as positive controls is proposed. The in vitro transcribed RNA could also be used as quality standards in the design of international proficiency studies. © 2013 Elsevier B.V.

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DO - 10.1016/j.jviromet.2013.07.016

M3 - Article

SP - 487

EP - 491

JO - Journal of Virological Methods

JF - Journal of Virological Methods

SN - 0166-0934

ER -