1. Cas-OPRAD: a one-pot RPA/PCR CRISPR/Cas12 assay for on-site Phytophthora root rot detection
- Author
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Zhiting Li, Wanzhen Feng, Zaobing Zhu, Shengdan Lu, Mingze Lin, Jiali Dong, Zhixin Wang, Fuxiu Liu, and Qinghe Chen
- Subjects
Phytophthora sojae ,one-pot RPA/PCR ,recombinase polymerase amplification ,CRISPR/Cas12a ,on-site detection ,visual detection ,Microbiology ,QR1-502 - Abstract
Phytophthora sojae is a devastating plant pathogen that causes soybean Phytophthora root rot worldwide. Early on-site and accurate detection of the causal pathogen is critical for successful management. In this study, we have developed a novel and specific one-pot RPA/PCR-CRISPR/Cas12 assay for on-site detection (Cas-OPRAD) of Phytophthora root rot (P. sojae). Compared to the traditional RPA/PCR detection methods, the Cas-OPRAD assay has significant detection performance. The Cas-OPRAD platform has excellent specificity to distinguish 33 P. sojae from closely related oomycetes or fungal species. The PCR-Cas12a assay had a consistent detection limit of 100 pg. μL−1, while the RPA-Cas12a assay achieved a detection limit of 10 pg. μL−1. Furthermore, the Cas-OPRAD assay was equipped with a lateral flow assay for on-site diagnosis and enabled the visual detection of P. sojae on the infected field soybean samples. This assay provides a simple, efficient, rapid (
- Published
- 2024
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