A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA Sequences
Adulteration of meat with carnivorous animals (such as cats, dogs, foxes, and minks) can cause ethical problems and lead to disease transmission; however, DNA quantitative methods for four carnivorous species in one tube reaction are still rare. In this study, a carnivore-specific nuclear DNA sequen...
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MDPI AG
2023-01-01
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| Series: | Foods |
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| Online Access: | https://www.mdpi.com/2304-8158/12/3/594 |
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| author | Wenjun Wang Tiean Wei Manna Shi Yu Han Yang Shen Xiang Zhou Bang Liu |
| author_facet | Wenjun Wang Tiean Wei Manna Shi Yu Han Yang Shen Xiang Zhou Bang Liu |
| author_sort | Wenjun Wang |
| collection | DOAJ |
| description | Adulteration of meat with carnivorous animals (such as cats, dogs, foxes, and minks) can cause ethical problems and lead to disease transmission; however, DNA quantitative methods for four carnivorous species in one tube reaction are still rare. In this study, a carnivore-specific nuclear DNA sequence that is conserved in carnivorous animals but has base differences within the sequence was used to design universal primers for its conserved region and corresponding species-specific probes for the hypervariable region. A novel universal primer multiplex real-time PCR (UP-M-rtPCR) approach was developed for the specific identification and quantitation of cat, dog, fox, and mink fractions in a single reaction, with a 0.05 ng absolute limit of detection (LOD) and 0.05% relative LOD. This approach simplifies the PCR system and improves the efficiency of simultaneous identification of multiple animal-derived ingredients in meat. UP-M-rtPCR showed good accuracy (0.48–7.04% relative deviation) and precision (1.42–13.78% relative standard deviation) for quantitative analysis of cat, dog, fox, and mink DNA as well as excellent applicability for the evaluation of meat samples. |
| first_indexed | 2024-03-11T09:45:15Z |
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| id | doaj.art-a2457939fb264bf0a0913d88877973fd |
| institution | Directory Open Access Journal |
| issn | 2304-8158 |
| language | English |
| last_indexed | 2024-03-11T09:45:15Z |
| publishDate | 2023-01-01 |
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| record_format | Article |
| series | Foods |
| spelling | doaj.art-a2457939fb264bf0a0913d88877973fd2023-11-16T16:41:34ZengMDPI AGFoods2304-81582023-01-0112359410.3390/foods12030594A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA SequencesWenjun Wang0Tiean Wei1Manna Shi2Yu Han3Yang Shen4Xiang Zhou5Bang Liu6Key Laboratory of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, Hongshan Laboratory, Huazhong Agricultural University, Wuhan 430070, ChinaKey Laboratory of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, Hongshan Laboratory, Huazhong Agricultural University, Wuhan 430070, ChinaKey Laboratory of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, Hongshan Laboratory, Huazhong Agricultural University, Wuhan 430070, ChinaKey Laboratory of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, Hongshan Laboratory, Huazhong Agricultural University, Wuhan 430070, ChinaKey Laboratory of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, Hongshan Laboratory, Huazhong Agricultural University, Wuhan 430070, ChinaKey Laboratory of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, Hongshan Laboratory, Huazhong Agricultural University, Wuhan 430070, ChinaKey Laboratory of Agricultural Animal Genetics, Breeding, and Reproduction of Ministry of Education, Hongshan Laboratory, Huazhong Agricultural University, Wuhan 430070, ChinaAdulteration of meat with carnivorous animals (such as cats, dogs, foxes, and minks) can cause ethical problems and lead to disease transmission; however, DNA quantitative methods for four carnivorous species in one tube reaction are still rare. In this study, a carnivore-specific nuclear DNA sequence that is conserved in carnivorous animals but has base differences within the sequence was used to design universal primers for its conserved region and corresponding species-specific probes for the hypervariable region. A novel universal primer multiplex real-time PCR (UP-M-rtPCR) approach was developed for the specific identification and quantitation of cat, dog, fox, and mink fractions in a single reaction, with a 0.05 ng absolute limit of detection (LOD) and 0.05% relative LOD. This approach simplifies the PCR system and improves the efficiency of simultaneous identification of multiple animal-derived ingredients in meat. UP-M-rtPCR showed good accuracy (0.48–7.04% relative deviation) and precision (1.42–13.78% relative standard deviation) for quantitative analysis of cat, dog, fox, and mink DNA as well as excellent applicability for the evaluation of meat samples.https://www.mdpi.com/2304-8158/12/3/594carnivorous animalsnuclear DNA target sequencesuniversal primers multiplex real-time PCR (UP-M-rtPCR)quantitative detectionmeat adulteration |
| spellingShingle | Wenjun Wang Tiean Wei Manna Shi Yu Han Yang Shen Xiang Zhou Bang Liu A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA Sequences Foods carnivorous animals nuclear DNA target sequences universal primers multiplex real-time PCR (UP-M-rtPCR) quantitative detection meat adulteration |
| title | A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA Sequences |
| title_full | A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA Sequences |
| title_fullStr | A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA Sequences |
| title_full_unstemmed | A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA Sequences |
| title_short | A Novel Universal Primer Multiplex Real-Time PCR (UP-M-rtPCR) Approach for Specific Identification and Quantitation of Cat, Dog, Fox, and Mink Fractions Using Nuclear DNA Sequences |
| title_sort | novel universal primer multiplex real time pcr up m rtpcr approach for specific identification and quantitation of cat dog fox and mink fractions using nuclear dna sequences |
| topic | carnivorous animals nuclear DNA target sequences universal primers multiplex real-time PCR (UP-M-rtPCR) quantitative detection meat adulteration |
| url | https://www.mdpi.com/2304-8158/12/3/594 |
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