18S Ribosomal RNA Evaluation as Preanalytical Quality Control for Animal DNA.
- Leonard CA Department of Pathobiology, Institute of Veterinary Pathology, University of Zurich, Winterthurerstrasse 268, 8057 Zurich, Switzerland; Center for Clinical Studies, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 260, 8057 Zurich, Switzerland.
- Meli ML Center for Clinical Studies, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 260, 8057 Zurich, Switzerland; Clinical Laboratory, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 260, 8057 Zurich, Switzerland.
- Novacco M Center for Clinical Studies, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 260, 8057 Zurich, Switzerland; Clinical Laboratory, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 260, 8057 Zurich, Switzerland.
- Borel N Department of Pathobiology, Institute of Veterinary Pathology, University of Zurich, Winterthurerstrasse 268, 8057 Zurich, Switzerland; Center for Clinical Studies, Vetsuisse Faculty, University of Zurich, Winterthurerstrasse 260, 8057 Zurich, Switzerland.
- 2016-09-28
Published in:
- BioMed research international. - 2016
General Biochemistry, Genetics and Molecular Biology
General Immunology and Microbiology
General Medicine
English
The 18S ribosomal RNA (rRNA) gene is present in all eukaryotic cells. In this study, we evaluated the use of this gene to verify the presence of PCR-amplifiable host (animal) DNA as an indicator of sufficient sample quality for quantitative real-time PCR (qPCR) analysis. We compared (i) samples from various animal species, tissues, and sample types, including swabs; (ii) multiple DNA extraction methods; and (iii) both fresh and formalin-fixed paraffin-embedded (FFPE) samples. Results showed that 18S ribosomal RNA gene amplification was possible from all tissue samples evaluated, including avian, reptile, and FFPE samples and most swab samples. A single swine rectal swab, which showed sufficient DNA quantity and the demonstrated lack of PCR inhibitors, nonetheless was negative by 18S qPCR. Such a sample specifically illustrates the improvement of determination of sample integrity afforded by inclusion of 18S rRNA gene qPCR analysis in addition to spectrophotometric analysis and the use of internal controls for PCR inhibition. Other possible applications for the described 18S rRNA qPCR are preselection of optimal tissue specimens for studies or preliminary screening of archived samples prior to acceptance for biobanking projects.
- Language
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- English
- Open access status
- gold
- Identifiers
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- DOI 10.1155/2016/6104323
- PMID 27672657
- Persistent URL
- https://sonar.ch/global/documents/18484
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