Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasitica
Foot and root rot caused by Phytophthora parasitica is a substantial threat to citrus cultivation, affecting both yield and quality. Thus, rapid and accurate detection of P. parasitica plays an important role in disease management. The aim of this study was to develop a simple diagnostic method to d...
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| Format: | Article |
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KeAi Communications Co., Ltd.
2021-10-01
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| Series: | Journal of Integrative Agriculture |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2095311920634591 |
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| author | Wei-yu CHEN Jia YU Heng XU Xin-yu LU Ting-ting DAI Yue-e TIAN Dan-yu SHEN Dao-long DOU |
| author_facet | Wei-yu CHEN Jia YU Heng XU Xin-yu LU Ting-ting DAI Yue-e TIAN Dan-yu SHEN Dao-long DOU |
| author_sort | Wei-yu CHEN |
| collection | DOAJ |
| description | Foot and root rot caused by Phytophthora parasitica is a substantial threat to citrus cultivation, affecting both yield and quality. Thus, rapid and accurate detection of P. parasitica plays an important role in disease management. The aim of this study was to develop a simple diagnostic method to detect P. parasitica infection by combining recombinase polymerase amplification and lateral flow strips (LF-RPA). To establish the LF-RPA assay of P. parasitica, the primers and probe designed based on the Ypt1 gene were tested for specificity to P. parasitica, which showed no cross-reactivity with DNAs of other related oomycete species. The LF-RPA assay detected the amount of genomic DNA of P. parasitica which was as low as 1 pg. To make the LF-RPA assay useful in low-resource settings, four simplified DNA extraction methods were compared, after which the LF-RPA assay was applied, with no specialized equipment, to analyze a diverse range of citrus tissues by using a simplified PEG-NaOH method for DNA extraction. This method was successful in detecting P. parasitica in infected plant samples within 30 min. Combining the LF-RPA assay and a simplified DNA extraction method could be a potential detection test for P. parasitica, especially in areas with limited resources. |
| format | Article |
| id | doaj-art-ef2386733bc2480eac4609e97585d0a2 |
| institution | Kabale University |
| issn | 2095-3119 |
| language | English |
| publishDate | 2021-10-01 |
| publisher | KeAi Communications Co., Ltd. |
| record_format | Article |
| series | Journal of Integrative Agriculture |
| spelling | doaj-art-ef2386733bc2480eac4609e97585d0a22025-08-20T03:57:55ZengKeAi Communications Co., Ltd.Journal of Integrative Agriculture2095-31192021-10-0120102696270510.1016/S2095-3119(20)63459-1Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasiticaWei-yu CHEN0Jia YU1Heng XU2Xin-yu LU3Ting-ting DAI4Yue-e TIAN5Dan-yu SHEN6Dao-long DOU7Nanjing Plant Protection and Quarantine Station, Nanjing 210036, P.R.ChinaDepartment of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, P.R.China; College of Chemical and Biological Engineering, Taiyuan University of Science and Technology, Taiyuan 030024, P.R.ChinaDepartment of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, P.R.ChinaDepartment of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, P.R.ChinaCollege of Forestry, Nanjing Forestry University, Nanjing 210037, P.R.ChinaDepartment of Plant Protection, Henan University of Science and Technology, Luoyang 471000, P.R.ChinaDepartment of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, P.R.China; Correspondence SHEN Dan-yu, Tel: +86-25-84396355Department of Plant Pathology, Nanjing Agricultural University, Nanjing 210095, P.R.China; Correspondence DOU Dao-long, Tel: +86-25-84396973Foot and root rot caused by Phytophthora parasitica is a substantial threat to citrus cultivation, affecting both yield and quality. Thus, rapid and accurate detection of P. parasitica plays an important role in disease management. The aim of this study was to develop a simple diagnostic method to detect P. parasitica infection by combining recombinase polymerase amplification and lateral flow strips (LF-RPA). To establish the LF-RPA assay of P. parasitica, the primers and probe designed based on the Ypt1 gene were tested for specificity to P. parasitica, which showed no cross-reactivity with DNAs of other related oomycete species. The LF-RPA assay detected the amount of genomic DNA of P. parasitica which was as low as 1 pg. To make the LF-RPA assay useful in low-resource settings, four simplified DNA extraction methods were compared, after which the LF-RPA assay was applied, with no specialized equipment, to analyze a diverse range of citrus tissues by using a simplified PEG-NaOH method for DNA extraction. This method was successful in detecting P. parasitica in infected plant samples within 30 min. Combining the LF-RPA assay and a simplified DNA extraction method could be a potential detection test for P. parasitica, especially in areas with limited resources.http://www.sciencedirect.com/science/article/pii/S2095311920634591citrus diseasePhytophthora parasiticarecombined polymerase amplificationequipment-free detection |
| spellingShingle | Wei-yu CHEN Jia YU Heng XU Xin-yu LU Ting-ting DAI Yue-e TIAN Dan-yu SHEN Dao-long DOU Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasitica Journal of Integrative Agriculture citrus disease Phytophthora parasitica recombined polymerase amplification equipment-free detection |
| title | Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasitica |
| title_full | Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasitica |
| title_fullStr | Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasitica |
| title_full_unstemmed | Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasitica |
| title_short | Combining simplified DNA extraction technology and recombinase polymerase amplification assay for rapid and equipment-free detection of citrus pathogen Phytophthora parasitica |
| title_sort | combining simplified dna extraction technology and recombinase polymerase amplification assay for rapid and equipment free detection of citrus pathogen phytophthora parasitica |
| topic | citrus disease Phytophthora parasitica recombined polymerase amplification equipment-free detection |
| url | http://www.sciencedirect.com/science/article/pii/S2095311920634591 |
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