The qPCR and loop mediated isothermal amplification for rapid detection of Ustilago tritici
- Subject Areas
- Agricultural Science, Biotechnology, Microbiology, Molecular Biology
- LAMP, qPCR, wheat loose smut, Ustilago tritici
- © 2019 Yan et al.
- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ Preprints) and either DOI or URL of the article must be cited.
- Cite this article
- 2019. The qPCR and loop mediated isothermal amplification for rapid detection of Ustilago tritici. PeerJ Preprints 7:e27751v1 https://doi.org/10.7287/peerj.preprints.27751v1
Wheat loose smut caused by Ustilago tritici a seed-borne disease, is difficult to control due to the expansion of wheat planting area and difficulty of pathogen detection. In this study, real-time fluorescence quantitative PCR (qPCR) and loop-mediated isothermal amplification (LAMP) assays were used to rapidly amplify the DNA of U. tritici. Five pairs primers for qPCR and two series primers for LAMP were designed. Firstly, the specify of primers were carried out by using the DNAs of U. tritici, Fusarium graminearum, Blumeria graminis, Rhizoctonia cerealis, Puccinia striiformis, Bipolaris sorokiniana, and Alternaria solani as templates. Then the amplification systems are optimized. Finally, the sensitivity of qPCR and LAMP assays were quantified. The results show that using the primers pairs Y430F/R, Y307F/R, Y755F/R and Y139F/R for qPCR, primers L-139 and L-988 for LAMP assay could be used for U. tritici detection. In the sensitivity test, the detection limit of qPCR assay is 10 pg μL-1 of genomic DNA, the detection limit of LAMP assay is 100 fg μL -1 . We successfully performed qPCR and LAMP assays on two wheat loose smut wheat samples, and confirmed sequenced U. tritici infection by subsequently sequencing. This paper established two methods for U. tritici detection, which could be used for wheat loose smut diagnose in lab and field.
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