Establishment of a nucleic acid assay for detection of Echinococcus granulosus based on recombinase-aided isothermal amplification assay
10.16250/j.32.1374.2020071
- VernacularTitle:基于重组酶介导等温扩增技术的细粒棘球绦虫核酸检测方法的建立
- Author:
Xin DING
1
;
Yan-Hong LIU
2
;
Bi-Xian NI
1
;
Xiao-Ting WANG
1
;
Xiang-Zhen XU
1
;
Qing-Jie YING
2
;
Yang DAI
1
;
Jun CAO
1
Author Information
1. National Health Commission Key Laboratory of Parasitic Disease Control and Prevention, Jiangsu Provincial Key Laboratory on Parasite and Vector Control Technology, Jiangsu Institute of Parasitic Diseases, Public Health Research Center, Jiangnan University, Wuxi 214064, China
2. Jiangsu Qitian Gene Technology Co., Ltd., China
- Publication Type:Journal Article
- Keywords:
Echinococcus granulosus;
Recombinase;
Isothermal amplification;
Fluorescent probe;
Nucleic acid detection;
Diagnostic performance
- From:
Chinese Journal of Schistosomiasis Control
2020;32(4):340-344
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a nucleic acid assay for detection of Echinococcus granulosus based on recombinase-aided isothermal amplification (RAA) assay. Methods The 12S rRNA gene of E. granulosus was selected as the target gene, and the specific primers and fluorescent probes for RAA assay were designed, screened and synthesized to establish a fluorescent RAA assay for detection of E. granulosus. The sensitivity of the fluorescent RAA assay was evaluated using different copy numbers of target gene sequence-contained recombinant plasmids and various concentrations of E. granulosus genomic DNA as templates, and the specificity of the fluorescent RAA assay was evaluated using the genomic DNA from E. granulosus, E. multilocularis, Schistosoma japonicum, S. mansoni, Ancylostoma duodenale, Clonorchis sinensis, Taenia saginata, Spirometra mansoni and Taenia solium as templates. Results A fluorescent RAA assay was successfully established for detection of E. granulosus, which achieved specific amplification of E. granulosus genomic DNA within 20 min at 39 ℃. The lowest detection limit of the fluorescent RAA assay was 10 copies/μL of recombinant plasmids and 0.1 ng/μL E. granulosus genomic DNA, which exhibited a high sensitivity, and the fluorescent RAA assay was all negative for the genomic DNA from E. multilocularis, S. japonicum, S. mansoni, A. duodenale, C. sinensis, T. saginata, Spirometra mansoni and T. solium, which exhibited a high specificity. In addition, this fluorescent RAA assay successfully detected genomic DNA from E. granulosus cysts. Conclusions A rapid, sensitive and specific fluorescent RAA assay is successfully established for nucleic acid detection of E. granulosus.
