RPA-PCR couple : an approach to expedite plant diagnostics and overcome PCR inhibitors
DNA extraction can be lengthy and sometimes ends up with amplification inhibitors. We present the potential of recombinase polymerase amplification (RPA) to replace plant DNA extraction. In our rapid 'RPA-PCR couple' concept, RPA is tuned to slower reaction kinetics to promote amplificatio...
| Veröffentlicht in: | BioTechniques. - 1988. - 69(2020), 4 vom: 18. Okt., Seite 270-280 |
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| Format: | Online-Aufsatz |
| Sprache: | English |
| Veröffentlicht: |
2020
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| Zugriff auf das übergeordnete Werk: | BioTechniques |
| Schlagworte: | Journal Article Research Support, Non-U.S. Gov't PCR inhibitors Phytophthora Phytophthora cactorum Phytophthora fragariae atp9-nad9 plant DNA extraction recombinase polymerase amplification Benzothiazoles mehr... |
| Zusammenfassung: | DNA extraction can be lengthy and sometimes ends up with amplification inhibitors. We present the potential of recombinase polymerase amplification (RPA) to replace plant DNA extraction. In our rapid 'RPA-PCR couple' concept, RPA is tuned to slower reaction kinetics to promote amplification of long targets. RPA primers amplify target and some flanking regions directly from simple plant macerates. Then PCR primers exponentially amplify the target directly from the RPA reaction. We present the coupling of RPA with conventional, TaqMan and SYBR Green PCR assays. We applied the concept to strawberry Phytophthora pathogens and the Phytophthora identification marker atp9-nad9. We found RPA-PCR couple specific, sensitive and reliable. The approach may also benefit other difficult samples such as food, feces and ancient samples |
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| Beschreibung: | Date Completed 09.08.2021 Date Revised 09.08.2021 published: Print-Electronic Citation Status MEDLINE |
| ISSN: | 1940-9818 |
| DOI: | 10.2144/btn-2020-0065 |