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A novel multiplex xMAP assay for generic detection of avian, fish, and ruminant DNA in feed and feedstuffs

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Abstract

The identification of animal species in feed and feedstuffs is important for detecting contamination and fraudulent replacement of animal components that might cause health and economic problems. A novel multiplex assay, based on xMAP technology and the generic detection of closely related species, was developed for the simultaneous differential detection of avian, fish, and ruminant DNA in products. Universal primers and probes specific to avian, fish, or ruminant species were designed to target a conserved mitochondrial DNA sequence in the 12S ribosomal RNA gene (rRNA). The assay specificity was validated using samples of 27 target and 10 nontarget animal species. The limits of detection of the purified DNA were determined to be 0.2 pg/μL–0.1 ng/μL by testing the meat samples of six species and four feedstuffs. The detection sensitivity of the experimental mixtures was demonstrated to be 0.01% (weight percentage). The assay’s suitability for practical application was evaluated by testing feed samples; unlabeled animal ingredients were detected in 32% of the 56 samples. The assay differentially detected the three targeted categories of animal species in less than 2 h, reflecting improvements in speed and efficiency. Based on these results, this novel multiplex xMAP assay provides a reliable and highly efficient technology for the routine detection of animal species in feed and other products for which this information is needed.

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Acknowledgments

We thank Hai-Yan Luo and Xin Tan for technical assistance.

Funding

This work was supported by the Science Research Program of Guangzhou, China (grant no. 201707010487) and the Science and Technology Research Project of the former Guangdong Entry-exit Inspection and Quarantine Bureau, China (grant no. 2018GDK04).

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Authors and Affiliations

  1. Technical Center, Guangzhou Customs District People’s Republic of China, Guangzhou, 510623, China

    Ru Chen, Ming-Zhu Mei, Yan-Yu Duan, Zhi-Ling Liu, Wen-Chuan Weng & Jing Yang

  2. Department of Genetics, National Research Institute for Family Planning, Beijing, 100081, China

    Xiao-Bo Gao

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  1. Ru Chen
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  2. Xiao-Bo Gao
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  3. Ming-Zhu Mei
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  4. Yan-Yu Duan
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  6. Wen-Chuan Weng
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  7. Jing Yang
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Correspondence to Ru Chen.

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Chen, R., Gao, XB., Mei, MZ. et al. A novel multiplex xMAP assay for generic detection of avian, fish, and ruminant DNA in feed and feedstuffs. Appl Microbiol Biotechnol 103, 4575–4584 (2019). https://doi.org/10.1007/s00253-019-09833-9

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  • DOI: https://doi.org/10.1007/s00253-019-09833-9

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