Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach
Peach bacterial spot caused by Xanthomonas arboricola pv. pruni (Xap) is a devastating disease worldwide and frequently causes massive economic losses. In recent years, it has become a pandemic outbreak in most peach production areas of China, especially on precocious peaches in the middle reach of...
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Frontiers Media S.A.
2021-11-01
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| Series: | Frontiers in Plant Science |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fpls.2021.740177/full |
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| author | Mei Luo Fan-Zhu Meng Qin Tan Wei-Xiao Yin Chao-Xi Luo Chao-Xi Luo |
| author_facet | Mei Luo Fan-Zhu Meng Qin Tan Wei-Xiao Yin Chao-Xi Luo Chao-Xi Luo |
| author_sort | Mei Luo |
| collection | DOAJ |
| description | Peach bacterial spot caused by Xanthomonas arboricola pv. pruni (Xap) is a devastating disease worldwide and frequently causes massive economic losses. In recent years, it has become a pandemic outbreak in most peach production areas of China, especially on precocious peaches in the middle reach of the Yangtze River. Rapid, user-friendly detection is extremely important to make the correct diagnosis and develop suitable control strategies. In this study, we described a recombinase polymerase amplification (RPA)/Cas12a-based system that combines RPA and CRISPR/Cas12a for Xap identification. A total of three crRNAs were designed to target a highly conserved ABC transporter ATP-binding protein-encoding gene ftsX to make specific detection of Xap. Results showed that crRNA 2 and crRNA 3 could get consistent detection for Xap. To realize the visualization of detection results, we additionally introduced FQ-reporter and FB-reporter. The developed method was highly sensitive and could detect as low as 10–18 M Xap gDNA with a mini-UV torch, corresponding to 1.63 copies/μl or 8.855 fg/μl gDNA of Xap, while with lateral flow strips, the sensitivity was 10–17 M. In addition, this method could specifically detect Xap from other closely related bacteria or pathogens associated with peach diseases. Furthermore, this method could make correct identification for Xap with crude DNA using NaOH-based extraction (3 min) directly from diseased peach samples. Considering that the developed method could get results within 2 h and could be performed at 37°C (body temperature), it is promising to be applied for Xap diagnosis and monitoring in fields. |
| first_indexed | 2024-12-19T17:42:45Z |
| format | Article |
| id | doaj.art-ac4f23fa477e4913a4b27062d620d162 |
| institution | Directory Open Access Journal |
| issn | 1664-462X |
| language | English |
| last_indexed | 2024-12-19T17:42:45Z |
| publishDate | 2021-11-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Plant Science |
| spelling | doaj.art-ac4f23fa477e4913a4b27062d620d1622022-12-21T20:12:09ZengFrontiers Media S.A.Frontiers in Plant Science1664-462X2021-11-011210.3389/fpls.2021.740177740177Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on PeachMei Luo0Fan-Zhu Meng1Qin Tan2Wei-Xiao Yin3Chao-Xi Luo4Chao-Xi Luo5Key Laboratory of Horticultural Plant Biology, Ministry of Education, Huazhong Agricultural University, Wuhan, ChinaHubei Key Laboratory of Plant Pathology, Huazhong Agricultural University, Wuhan, ChinaKey Laboratory of Horticultural Plant Biology, Ministry of Education, Huazhong Agricultural University, Wuhan, ChinaHubei Key Laboratory of Plant Pathology, Huazhong Agricultural University, Wuhan, ChinaKey Laboratory of Horticultural Plant Biology, Ministry of Education, Huazhong Agricultural University, Wuhan, ChinaHubei Key Laboratory of Plant Pathology, Huazhong Agricultural University, Wuhan, ChinaPeach bacterial spot caused by Xanthomonas arboricola pv. pruni (Xap) is a devastating disease worldwide and frequently causes massive economic losses. In recent years, it has become a pandemic outbreak in most peach production areas of China, especially on precocious peaches in the middle reach of the Yangtze River. Rapid, user-friendly detection is extremely important to make the correct diagnosis and develop suitable control strategies. In this study, we described a recombinase polymerase amplification (RPA)/Cas12a-based system that combines RPA and CRISPR/Cas12a for Xap identification. A total of three crRNAs were designed to target a highly conserved ABC transporter ATP-binding protein-encoding gene ftsX to make specific detection of Xap. Results showed that crRNA 2 and crRNA 3 could get consistent detection for Xap. To realize the visualization of detection results, we additionally introduced FQ-reporter and FB-reporter. The developed method was highly sensitive and could detect as low as 10–18 M Xap gDNA with a mini-UV torch, corresponding to 1.63 copies/μl or 8.855 fg/μl gDNA of Xap, while with lateral flow strips, the sensitivity was 10–17 M. In addition, this method could specifically detect Xap from other closely related bacteria or pathogens associated with peach diseases. Furthermore, this method could make correct identification for Xap with crude DNA using NaOH-based extraction (3 min) directly from diseased peach samples. Considering that the developed method could get results within 2 h and could be performed at 37°C (body temperature), it is promising to be applied for Xap diagnosis and monitoring in fields.https://www.frontiersin.org/articles/10.3389/fpls.2021.740177/fullCRISPR/Cas12aRPAXanthomonas arboricola pv. prunipeachbacterial spot |
| spellingShingle | Mei Luo Fan-Zhu Meng Qin Tan Wei-Xiao Yin Chao-Xi Luo Chao-Xi Luo Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach Frontiers in Plant Science CRISPR/Cas12a RPA Xanthomonas arboricola pv. pruni peach bacterial spot |
| title | Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach |
| title_full | Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach |
| title_fullStr | Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach |
| title_full_unstemmed | Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach |
| title_short | Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach |
| title_sort | recombinase polymerase amplification cas12a based identification of xanthomonas arboricola pv pruni on peach |
| topic | CRISPR/Cas12a RPA Xanthomonas arboricola pv. pruni peach bacterial spot |
| url | https://www.frontiersin.org/articles/10.3389/fpls.2021.740177/full |
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