Development of a novel recombinant encapsidated RNA particle: evaluation as an internal control for diagnostic RT-PCR

Research output: Contribution to journalJournal articleResearchpeer-review

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Development of a novel recombinant encapsidated RNA particle : evaluation as an internal control for diagnostic RT-PCR. / King, Donald P; Montague, Nick; Ebert, Katja; Reid, Scott M; Dukes, Juliet P; Schädlich, Lysann; Belsham, Graham J; Lomonossoff, George P.

In: Journal of Virological Methods, Vol. 146, No. 1-2, 12.2007, p. 218-25.

Research output: Contribution to journalJournal articleResearchpeer-review

Harvard

King, DP, Montague, N, Ebert, K, Reid, SM, Dukes, JP, Schädlich, L, Belsham, GJ & Lomonossoff, GP 2007, 'Development of a novel recombinant encapsidated RNA particle: evaluation as an internal control for diagnostic RT-PCR', Journal of Virological Methods, vol. 146, no. 1-2, pp. 218-25. https://doi.org/10.1016/j.jviromet.2007.07.002

APA

King, D. P., Montague, N., Ebert, K., Reid, S. M., Dukes, J. P., Schädlich, L., Belsham, G. J., & Lomonossoff, G. P. (2007). Development of a novel recombinant encapsidated RNA particle: evaluation as an internal control for diagnostic RT-PCR. Journal of Virological Methods, 146(1-2), 218-25. https://doi.org/10.1016/j.jviromet.2007.07.002

Vancouver

King DP, Montague N, Ebert K, Reid SM, Dukes JP, Schädlich L et al. Development of a novel recombinant encapsidated RNA particle: evaluation as an internal control for diagnostic RT-PCR. Journal of Virological Methods. 2007 Dec;146(1-2):218-25. https://doi.org/10.1016/j.jviromet.2007.07.002

Author

King, Donald P ; Montague, Nick ; Ebert, Katja ; Reid, Scott M ; Dukes, Juliet P ; Schädlich, Lysann ; Belsham, Graham J ; Lomonossoff, George P. / Development of a novel recombinant encapsidated RNA particle : evaluation as an internal control for diagnostic RT-PCR. In: Journal of Virological Methods. 2007 ; Vol. 146, No. 1-2. pp. 218-25.

Bibtex

@article{44d234c060cc48fdac58db852f782802,
title = "Development of a novel recombinant encapsidated RNA particle: evaluation as an internal control for diagnostic RT-PCR",
abstract = "This report describes the generation of novel encapsidated RNA particles and their evaluation as in-tube internal controls in diagnostic real-time reverse-transcription PCR (rRT-PCR) assays for the detection of RNA viruses. A cassette containing sequences of 2 diagnostic primer sets for foot-and-mouth disease virus (FMDV) and a set for swine vesicular disease virus (SVDV) was engineered into a full-length cDNA clone containing the RNA-2 segment of Cowpea Mosaic Virus (CPMV). After co-inoculation with a plasmid that expressed CPMV RNA-1, recombinant virus particles were rescued from cowpea plants (Vigna unguiculata). RNA contained in these particles was amplified in diagnostic rRT-PCR assays used for detection of FMDV and SVDV. Amplification of these internal controls was used to confirm that rRT-PCR inhibitors were absent from clinical samples, thereby verifying negative assay results. The recombinant CPMVs did not reduce the analytical sensitivity of the rRT-PCRs when amplification of the insert was performed in the same tube as the diagnostic target. This system provides an attractive solution to the production of internal controls for rRT-PCR assays since CPMV grows to high yields in plants, the particles are thermostable, RNase resistant and simple purification of RNA-2 containing capsids yields a preparation which is non-infectious.",
keywords = "Animals, Comovirus/genetics, Enterovirus B, Human/isolation & purification, Foot-and-Mouth Disease/virology, Foot-and-Mouth Disease Virus/isolation & purification, Genetic Vectors, RNA/genetics, Reverse Transcriptase Polymerase Chain Reaction/methods, Sensitivity and Specificity, Swine/virology, Swine Vesicular Disease/virology",
author = "King, {Donald P} and Nick Montague and Katja Ebert and Reid, {Scott M} and Dukes, {Juliet P} and Lysann Sch{\"a}dlich and Belsham, {Graham J} and Lomonossoff, {George P}",
year = "2007",
month = dec,
doi = "10.1016/j.jviromet.2007.07.002",
language = "English",
volume = "146",
pages = "218--25",
journal = "Journal of Virological Methods",
issn = "0166-0934",
publisher = "Elsevier",
number = "1-2",

}

RIS

TY - JOUR

T1 - Development of a novel recombinant encapsidated RNA particle

T2 - evaluation as an internal control for diagnostic RT-PCR

AU - King, Donald P

AU - Montague, Nick

AU - Ebert, Katja

AU - Reid, Scott M

AU - Dukes, Juliet P

AU - Schädlich, Lysann

AU - Belsham, Graham J

AU - Lomonossoff, George P

PY - 2007/12

Y1 - 2007/12

N2 - This report describes the generation of novel encapsidated RNA particles and their evaluation as in-tube internal controls in diagnostic real-time reverse-transcription PCR (rRT-PCR) assays for the detection of RNA viruses. A cassette containing sequences of 2 diagnostic primer sets for foot-and-mouth disease virus (FMDV) and a set for swine vesicular disease virus (SVDV) was engineered into a full-length cDNA clone containing the RNA-2 segment of Cowpea Mosaic Virus (CPMV). After co-inoculation with a plasmid that expressed CPMV RNA-1, recombinant virus particles were rescued from cowpea plants (Vigna unguiculata). RNA contained in these particles was amplified in diagnostic rRT-PCR assays used for detection of FMDV and SVDV. Amplification of these internal controls was used to confirm that rRT-PCR inhibitors were absent from clinical samples, thereby verifying negative assay results. The recombinant CPMVs did not reduce the analytical sensitivity of the rRT-PCRs when amplification of the insert was performed in the same tube as the diagnostic target. This system provides an attractive solution to the production of internal controls for rRT-PCR assays since CPMV grows to high yields in plants, the particles are thermostable, RNase resistant and simple purification of RNA-2 containing capsids yields a preparation which is non-infectious.

AB - This report describes the generation of novel encapsidated RNA particles and their evaluation as in-tube internal controls in diagnostic real-time reverse-transcription PCR (rRT-PCR) assays for the detection of RNA viruses. A cassette containing sequences of 2 diagnostic primer sets for foot-and-mouth disease virus (FMDV) and a set for swine vesicular disease virus (SVDV) was engineered into a full-length cDNA clone containing the RNA-2 segment of Cowpea Mosaic Virus (CPMV). After co-inoculation with a plasmid that expressed CPMV RNA-1, recombinant virus particles were rescued from cowpea plants (Vigna unguiculata). RNA contained in these particles was amplified in diagnostic rRT-PCR assays used for detection of FMDV and SVDV. Amplification of these internal controls was used to confirm that rRT-PCR inhibitors were absent from clinical samples, thereby verifying negative assay results. The recombinant CPMVs did not reduce the analytical sensitivity of the rRT-PCRs when amplification of the insert was performed in the same tube as the diagnostic target. This system provides an attractive solution to the production of internal controls for rRT-PCR assays since CPMV grows to high yields in plants, the particles are thermostable, RNase resistant and simple purification of RNA-2 containing capsids yields a preparation which is non-infectious.

KW - Animals

KW - Comovirus/genetics

KW - Enterovirus B, Human/isolation & purification

KW - Foot-and-Mouth Disease/virology

KW - Foot-and-Mouth Disease Virus/isolation & purification

KW - Genetic Vectors

KW - RNA/genetics

KW - Reverse Transcriptase Polymerase Chain Reaction/methods

KW - Sensitivity and Specificity

KW - Swine/virology

KW - Swine Vesicular Disease/virology

U2 - 10.1016/j.jviromet.2007.07.002

DO - 10.1016/j.jviromet.2007.07.002

M3 - Journal article

C2 - 17727966

VL - 146

SP - 218

EP - 225

JO - Journal of Virological Methods

JF - Journal of Virological Methods

SN - 0166-0934

IS - 1-2

ER -

ID: 257918608