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Simultaneous detection of three wheat pathogenic fungal species by multiplex PCR

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Abstract

Wheat stem rust, wheat leaf rust, and wheat powdery mildew are three of the most economically damaging fungal diseases of wheat in China, caused by Puccinia graminis f. sp. tritici, Puccinia triticina, and Blumeria graminis f. sp. tritici, respectively. In this study, single-step multiplex polymerase chain reaction (PCR) was developed for the simultaneous detection of the three pathogens. Three sets of specific primers were selected, followed by optimization of the annealing temperature and the dNTPs, Taq DNA polymerase, and primer concentrations for multiplex PCR to establish a method for the simultaneous detection of P. graminis f. sp. tritici, P. triticina, and B. graminis f. sp. tritici at an early stage of infection. The specificity and sensitivity of the multiplex PCR were tested. We observed three specific bands of 395 bp, 151 bp, and 464 bp on amplification with a three-reaction system. The detection sensitivity of the multiplex PCR was 1 ng DNA for P. graminis f. sp. tritici, 10 pg DNA for P. triticina, and 10 pg DNA for B. graminis f. sp. tritici. This study pioneered the use of the triplex PCR system for simultaneous detection of P. graminis f. sp. tritici, P. triticina, and B. graminis f. sp. tritici in infected plant tissues. This novel PCR-based method provides a simple and rapid method for detecting various pathogens in wheat, which will assist in the simultaneous diagnosis and monitoring of multiple plant diseases.

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Acknowledgments

This study was supported by the National Key Basic Research Program of China (2013CB127701), the National Natural Science Foundation of China (31171829), and the Special Fund for Agro-scientific Research in the Public Interest (201303016).

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Authors and Affiliations

  1. College of Plant Protection, Shenyang Agricultural University, Shenyang, Liaoning, 110866, People’s Republic of China

    Si Chen, Yuan-yin Cao, Tian-ya Li & Xian-xin Wu

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  1. Si Chen
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  2. Yuan-yin Cao
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  3. Tian-ya Li
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  4. Xian-xin Wu
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Correspondence to Yuan-yin Cao.

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Chen, S., Cao, Yy., Li, Ty. et al. Simultaneous detection of three wheat pathogenic fungal species by multiplex PCR. Phytoparasitica 43, 449–460 (2015). https://doi.org/10.1007/s12600-014-0442-1

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