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Establishment and Application of a Multiplex PCR Assay for Detection of Sclerotium rolfsii, Lasiodiplodia theobromae, and Fusarium oxysporum in Peanut

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Abstract

Southern blight, stem rot, and root rot are serious soil-borne fungal diseases of peanut, which are caused by Sclerotium rolfsii, Lasiodiplodia theobromae, and Fusarium oxysporum, respectively. These diseases are difficult to be diagnosed in early stage of infection, causing the optimal treatment period was often missed. Therefore, establishing a rapid detection system is of great significance for early prevention of peanut soil-borne fungal diseases. Here, we have invented a multiplex PCR detection system to detect fungal pathogens of peanut southern blight, stem rot, and root rot at the same time. The quarantine fungal pathogen primer pairs were amplified to the specific number of base pairs in each of the following fungal pathogens: 1005-bp (F. oxysporum), 238-bp (L. theobromae), and 638-bp (S. rolfsii). The detection limit for the single and multiplex PCR primer sets was 1 ng of template DNA under in vitro conditions. Amplification of fungi of non-target species yielded no non-specific products. The validation showed that the multiplex PCR could effectively detect single and mixed infections in field samples. Overview, this study proved that this mPCR assay was a rapid, reliable, and simple tool for the simultaneous detection of three important peanut soil-borne diseases, which facilitated prompt treatment and prevention of peanut root diseases.

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Data Availability

The data that support the findings of this study are available on request from the corresponding author, upon reasonable request.

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Acknowledgements

This study was supported by the National Natural Sciences Foundation of China (Grant No. 32202266), the Shandong Modern Agricultural Industry Technology System (Grant No. SDAIT-04-08), the Outstanding Youth Foundation of Shandong Province (Grant No. ZR2021YQ20), and the Key Research and Development Program of Shandong Province (Grant No. 2019JZZY020608).

Funding

Funding was provided by the National Natural Sciences Foundation of China (Grant No. 32202266), the Shandong Modern Agricultural Industry Technology System (Grant No. SDAIT-04-08), the Outstanding Youth Foundation of Shandong Province (Grant No. ZR2021YQ20), and the Key Research and Development Program of Shandong Province (Grant No. 2019JZZY020608).

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  1. Chao Han and Aixin Liu have contributed equally to this work.

Authors and Affiliations

  1. Shandong Provincial Key Laboratory of Agricultural Microbiology, College of Plant Protection, Shandong Agricultural University, Tai’an, 271018, China

    Jin Wang, Xue Li, Xueying Sun, Xuelin Huo, Meiqi Li, Chao Han & Aixin Liu

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  1. Jin Wang
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Contributions

AXL, XL, and JW conceived and designed research. XL, JW, XLH, and MQL conducted experiments. XYS and CH analyzed data. JW and CH wrote the manuscript. All authors read and approved the manuscript.

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Wang, J., Li, X., Sun, X. et al. Establishment and Application of a Multiplex PCR Assay for Detection of Sclerotium rolfsii, Lasiodiplodia theobromae, and Fusarium oxysporum in Peanut. Mol Biotechnol 65, 1369–1377 (2023). https://doi.org/10.1007/s12033-022-00647-1

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