Development of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquids
Abstract Background Nowadays, there is an increasing demand for fast and reliable plant biomolecular analyses. Conventional methods for the isolation of nucleic acids are time-consuming and require multiple and often non-automatable steps to remove cellular interferences, with consequence that sampl...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2019-03-01
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| Series: | Plant Methods |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s13007-019-0408-x |
| _version_ | 1811233825710145536 |
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| author | Arianna Marengo Cecilia Cagliero Barbara Sgorbini Jared L. Anderson Miranda N. Emaus Carlo Bicchi Cinzia M. Bertea Patrizia Rubiolo |
| author_facet | Arianna Marengo Cecilia Cagliero Barbara Sgorbini Jared L. Anderson Miranda N. Emaus Carlo Bicchi Cinzia M. Bertea Patrizia Rubiolo |
| author_sort | Arianna Marengo |
| collection | DOAJ |
| description | Abstract Background Nowadays, there is an increasing demand for fast and reliable plant biomolecular analyses. Conventional methods for the isolation of nucleic acids are time-consuming and require multiple and often non-automatable steps to remove cellular interferences, with consequence that sample preparation is the major bottleneck in the bioanalytical workflow. New opportunities have been created by the use of magnetic ionic liquids (MILs) thanks to their affinity for nucleic acids. Results In the present study, a MIL-based magnet-assisted dispersive liquid–liquid microextraction (maDLLME) method was optimized for the extraction of genomic DNA from Arabidopsis thaliana (L.) Heynh leaves. MILs containing different metal centers were tested and the extraction method was optimized in terms of MIL volume and extraction time for purified DNA and crude lysates. The proposed approach yielded good extraction efficiency and is compatible with both quantitative analysis through fluorimetric-based detection and qualitative analysis as PCR amplification of multi and single locus genes. The protocol was successfully applied to a set of plant species and tissues. Conclusions The developed MIL-based maDLLME approach exhibits good enrichment of nucleic acids for extraction of template suitable for targeted PCR; it is very fast, sustainable and potentially automatable thereby representing a powerful tool for screening plants rapidly using DNA-based methods. |
| first_indexed | 2024-04-12T11:25:49Z |
| format | Article |
| id | doaj.art-d2bdd72df18247d3981125a8c8843c91 |
| institution | Directory Open Access Journal |
| issn | 1746-4811 |
| language | English |
| last_indexed | 2024-04-12T11:25:49Z |
| publishDate | 2019-03-01 |
| publisher | BMC |
| record_format | Article |
| series | Plant Methods |
| spelling | doaj.art-d2bdd72df18247d3981125a8c8843c912022-12-22T03:35:14ZengBMCPlant Methods1746-48112019-03-0115111110.1186/s13007-019-0408-xDevelopment of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquidsArianna Marengo0Cecilia Cagliero1Barbara Sgorbini2Jared L. Anderson3Miranda N. Emaus4Carlo Bicchi5Cinzia M. Bertea6Patrizia Rubiolo7Dipartimento di Scienza e Tecnologia del Farmaco, Università di TorinoDipartimento di Scienza e Tecnologia del Farmaco, Università di TorinoDipartimento di Scienza e Tecnologia del Farmaco, Università di TorinoDepartment of Chemistry, Iowa State UniversityDepartment of Chemistry, Iowa State UniversityDipartimento di Scienza e Tecnologia del Farmaco, Università di TorinoDipartimento di Scienze della Vita e Biologia dei Sistemi, Unità di Fisiologia Vegetale, Università di TorinoDipartimento di Scienza e Tecnologia del Farmaco, Università di TorinoAbstract Background Nowadays, there is an increasing demand for fast and reliable plant biomolecular analyses. Conventional methods for the isolation of nucleic acids are time-consuming and require multiple and often non-automatable steps to remove cellular interferences, with consequence that sample preparation is the major bottleneck in the bioanalytical workflow. New opportunities have been created by the use of magnetic ionic liquids (MILs) thanks to their affinity for nucleic acids. Results In the present study, a MIL-based magnet-assisted dispersive liquid–liquid microextraction (maDLLME) method was optimized for the extraction of genomic DNA from Arabidopsis thaliana (L.) Heynh leaves. MILs containing different metal centers were tested and the extraction method was optimized in terms of MIL volume and extraction time for purified DNA and crude lysates. The proposed approach yielded good extraction efficiency and is compatible with both quantitative analysis through fluorimetric-based detection and qualitative analysis as PCR amplification of multi and single locus genes. The protocol was successfully applied to a set of plant species and tissues. Conclusions The developed MIL-based maDLLME approach exhibits good enrichment of nucleic acids for extraction of template suitable for targeted PCR; it is very fast, sustainable and potentially automatable thereby representing a powerful tool for screening plants rapidly using DNA-based methods.http://link.springer.com/article/10.1186/s13007-019-0408-xDNA isolationMagnetic ionic liquidsArabidopsis thaliana (L.) Heynh.DNA barcoding |
| spellingShingle | Arianna Marengo Cecilia Cagliero Barbara Sgorbini Jared L. Anderson Miranda N. Emaus Carlo Bicchi Cinzia M. Bertea Patrizia Rubiolo Development of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquids Plant Methods DNA isolation Magnetic ionic liquids Arabidopsis thaliana (L.) Heynh. DNA barcoding |
| title | Development of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquids |
| title_full | Development of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquids |
| title_fullStr | Development of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquids |
| title_full_unstemmed | Development of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquids |
| title_short | Development of an innovative and sustainable one-step method for rapid plant DNA isolation for targeted PCR using magnetic ionic liquids |
| title_sort | development of an innovative and sustainable one step method for rapid plant dna isolation for targeted pcr using magnetic ionic liquids |
| topic | DNA isolation Magnetic ionic liquids Arabidopsis thaliana (L.) Heynh. DNA barcoding |
| url | http://link.springer.com/article/10.1186/s13007-019-0408-x |
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