Engineered DNA Polymerase Improves PCR Results for Plastid DNA
Premise of the study: Secondary metabolites often inhibit PCR and sequencing reactions in extractions from plant material, especially from silica-dried and herbarium material. A DNA polymerase that is tolerant to inhibitors improves PCR results. Methods and Results: A novel DNA amplification syst...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Wiley
2013-02-01
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| Series: | Applications in Plant Sciences |
| Subjects: | |
| Online Access: | http://www.bioone.org/doi/full/10.3732/apps.1200519 |
| _version_ | 1819041962200662016 |
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| author | Melanie Schori Maryke Appel Alexarae Kitko Allan M. Showalter |
| author_facet | Melanie Schori Maryke Appel Alexarae Kitko Allan M. Showalter |
| author_sort | Melanie Schori |
| collection | DOAJ |
| description | Premise of the study: Secondary metabolites often inhibit PCR and sequencing reactions in extractions from plant material, especially from silica-dried and herbarium material. A DNA polymerase that is tolerant to inhibitors improves PCR results.
Methods and Results: A novel DNA amplification system, including a DNA polymerase engineered via directed evolution for improved tolerance to common plant-derived PCR inhibitors, was evaluated and PCR parameters optimized for three species. An additional 31 species were then tested with the engineered enzyme and optimized protocol, as well as with regular <i>Taq</i> polymerase.
Conclusions: PCR products and high-quality sequence data were obtained for 96% of samples for <i>rbcL</i> and 79% for <i>matK</i>, compared to 29% and 21% with regular <i>Taq</i> polymerase. |
| first_indexed | 2024-12-21T09:33:19Z |
| format | Article |
| id | doaj.art-3240f71a913d4371a8b198c330b2f58a |
| institution | Directory Open Access Journal |
| issn | 2168-0450 |
| language | English |
| last_indexed | 2024-12-21T09:33:19Z |
| publishDate | 2013-02-01 |
| publisher | Wiley |
| record_format | Article |
| series | Applications in Plant Sciences |
| spelling | doaj.art-3240f71a913d4371a8b198c330b2f58a2022-12-21T19:08:41ZengWileyApplications in Plant Sciences2168-04502013-02-0112120051910.3732/apps.1200519Engineered DNA Polymerase Improves PCR Results for Plastid DNAMelanie Schori0Maryke Appel1Alexarae Kitko2Allan M. Showalter3Department of Environmental and Plant Biology, Molecular and Cell Biology Program, Porter Hall 315, Ohio University, Athens, Ohio 45701 USAKapa Biosystems, 600 W. Cummings Park, Suite 2250, Woburn, Massachusetts 01801 USADepartment of Environmental and Plant Biology, Molecular and Cell Biology Program, Porter Hall 315, Ohio University, Athens, Ohio 45701 USA; 5833 Stearns Road, North Olmsted, Ohio 44070 USADepartment of Environmental and Plant Biology, Molecular and Cell Biology Program, Porter Hall 315, Ohio University, Athens, Ohio 45701 USAPremise of the study: Secondary metabolites often inhibit PCR and sequencing reactions in extractions from plant material, especially from silica-dried and herbarium material. A DNA polymerase that is tolerant to inhibitors improves PCR results. Methods and Results: A novel DNA amplification system, including a DNA polymerase engineered via directed evolution for improved tolerance to common plant-derived PCR inhibitors, was evaluated and PCR parameters optimized for three species. An additional 31 species were then tested with the engineered enzyme and optimized protocol, as well as with regular <i>Taq</i> polymerase. Conclusions: PCR products and high-quality sequence data were obtained for 96% of samples for <i>rbcL</i> and 79% for <i>matK</i>, compared to 29% and 21% with regular <i>Taq</i> polymerase.http://www.bioone.org/doi/full/10.3732/apps.1200519directed evolutionengineered KAPA3G DNA PolymerasematKPCR inhibitionrbcL. |
| spellingShingle | Melanie Schori Maryke Appel Alexarae Kitko Allan M. Showalter Engineered DNA Polymerase Improves PCR Results for Plastid DNA Applications in Plant Sciences directed evolution engineered KAPA3G DNA Polymerase matK PCR inhibition rbcL. |
| title | Engineered DNA Polymerase Improves PCR Results for Plastid DNA |
| title_full | Engineered DNA Polymerase Improves PCR Results for Plastid DNA |
| title_fullStr | Engineered DNA Polymerase Improves PCR Results for Plastid DNA |
| title_full_unstemmed | Engineered DNA Polymerase Improves PCR Results for Plastid DNA |
| title_short | Engineered DNA Polymerase Improves PCR Results for Plastid DNA |
| title_sort | engineered dna polymerase improves pcr results for plastid dna |
| topic | directed evolution engineered KAPA3G DNA Polymerase matK PCR inhibition rbcL. |
| url | http://www.bioone.org/doi/full/10.3732/apps.1200519 |
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