Medizinische Universität Graz - Research portal
Selected Publication:
Stöcher, M; Leb, V; Berg, J.
A convenient approach to the generation of multiple internal control DNA for a panel of real-time PCR assays.
J Virol Methods. 2003; 108(1):1-8
Doi: 10.1016/S0166-0934(02)00266-5
Web of Science
PubMed
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- Leading authors Med Uni Graz
- Berg Jörg
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- Abstract:
- Real-time polymerase chain reaction (PCR) assays allow convenient detection and quantitation of virus-derived nucleic acids in clinical specimens. When specimens are assayed for the presence of virus-derived nucleic acids against external standards, sample adequacy is not monitored. This can be achieved by using internal controls that are co-amplified with the virus-specific DNA in competitive PCR. Each of the various real-time PCR assays in a routine clinical laboratory requires its specific internal control. In order to complement a panel of virus-specific real-time PCR assays with internal controls, a convenient approach is described to generate the several internal controls within single DNA fragment. By applying composite primer technology, PCR primer sequences used in real-time PCR assays were added in 5' and 3' of a stretch of heterologous DNA during consecutive preparative PCRs. The heterologous DNA was used for internal control specific detection by e.g. FRET-hybridisation probes. The presented example of such a multiple internal control DNA contained five internal controls for five competitive LightCycler-PCR assays. All five PCR products derived from the multiple internal control DNA were detected with a single pair of specific FRET-hybridisation probes. The example described proved useful in real-time PCR assays specific for the detection of EBV-, CMV-, VZV- HSV-, and HBV-DNA on the LightCycler instrument. This methodology should enable laboratories to conveniently complement their panel of existing real-time PCR assays with a single multiple internal control DNA.
- Find related publications in this database (using NLM MeSH Indexing)
- Base Sequence -
- DNA Primers - genetics
- DNA, Viral - genetics DNA, Viral - standards
- Humans -
- Polymerase Chain Reaction - methods Polymerase Chain Reaction - standards
- Quality Control -
- Reference Standards -
- Virology - methods Virology - standards
- Find related publications in this database (Keywords)
- real-time PCR
- LightCycler
- internal control
- virus-derived nucleic acid