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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Autores principales:	Mei Luo, Fan-Zhu Meng, Qin Tan, Wei-Xiao Yin, Chao-Xi Luo
Formato:	article
Lenguaje:	EN
Publicado:	Frontiers Media S.A. 2021
Materias:	CRISPR/Cas12a RPA Xanthomonas arboricola pv. pruni peach bacterial spot Plant culture SB1-1110
Acceso en línea:	https://doaj.org/article/ac4f23fa477e4913a4b27062d620d162
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spelling	oai:doaj.org-article:ac4f23fa477e4913a4b27062d620d1622021-11-30T13:52:57ZRecombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach1664-462X10.3389/fpls.2021.740177https://doaj.org/article/ac4f23fa477e4913a4b27062d620d1622021-11-01T00:00:00Zhttps://www.frontiersin.org/articles/10.3389/fpls.2021.740177/fullhttps://doaj.org/toc/1664-462XPeach 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.Mei LuoFan-Zhu MengQin TanWei-Xiao YinChao-Xi LuoChao-Xi LuoFrontiers Media S.A.articleCRISPR/Cas12aRPAXanthomonas arboricola pv. prunipeachbacterial spotPlant cultureSB1-1110ENFrontiers in Plant Science, Vol 12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	CRISPR/Cas12a RPA Xanthomonas arboricola pv. pruni peach bacterial spot Plant culture SB1-1110
spellingShingle	CRISPR/Cas12a RPA Xanthomonas arboricola pv. pruni peach bacterial spot Plant culture SB1-1110 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
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.
format	article
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
title	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_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_sort	recombinase polymerase amplification/cas12a-based identification of xanthomonas arboricola pv. pruni on peach
publisher	Frontiers Media S.A.
publishDate	2021
url	https://doaj.org/article/ac4f23fa477e4913a4b27062d620d162
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Recombinase Polymerase Amplification/Cas12a-Based Identification of Xanthomonas arboricola pv. pruni on Peach

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