Recognizing false positives: synthetic oligonucleotide controls for environmental DNA surveillance
| Autor: | Chris C. Wilson, Kristyne M. Wozney, Caleigh M. Smith |
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| Rok vydání: | 2015 |
| Předmět: |
0106 biological sciences
0301 basic medicine Gel electrophoresis Oligonucleotide Ecological Modeling Biology 010603 evolutionary biology 01 natural sciences Molecular biology DNA sequencing Insert (molecular biology) 03 medical and health sciences Restriction enzyme 030104 developmental biology Real-time polymerase chain reaction False positive paradox Environmental DNA Ecology Evolution Behavior and Systematics |
| Zdroj: | Methods in Ecology and Evolution. 7:23-29 |
| ISSN: | 2041-210X |
| DOI: | 10.1111/2041-210x.12452 |
| Popis: | Summary Environmental DNA (eDNA) is increasingly used for surveillance and detection of species of interest in aquatic and soil samples. A significant risk associated with eDNA methods is potential false-positive results due to laboratory contamination. To minimize and quantify this risk, we designed and validated a set of synthetic oligonucleotides for use as species-specific positive PCR controls for several high-profile aquatic invasive species. The controls consist of species-specific sequences for the species of interest, with the addition of a synthetic insert containing recognition sites for several restriction enzymes. Following PCR, the presence of the synthetic insert can be detected using gel electrophoresis, restriction enzyme digests or DNA sequencing. For quantitative PCR (qPCR), false positives in environmental samples can also be detected using a fluorescent probe designed to detect the synthetic insert. The generation of synthetic controls is a cost-effective, reproducible method that increases the power and reliability of eDNA testing by eliminating misinterpretation of false-positive results from laboratory contamination. |
| Databáze: | OpenAIRE |
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