Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal Authentication
Point-of-care diagnostic methods for animal species determination are critical for rapid, simple, and accurate enforcement of food labelling. PCR is the most common method for species identification. However, the requirement of using a thermal cycler created drawbacks for the PCR application, partic...
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| Format: | Article |
| Language: | English |
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MDPI AG
2022-11-01
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| Series: | Molecules |
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| Online Access: | https://www.mdpi.com/1420-3049/27/23/8122 |
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| author | Mohd Hazim Mohd Yusop Mohd Fadzelly Abu Bakar Kamarul Rahim Kamarudin Nur Fadhilah Khairil Mokhtar Mohd Abd Motalib Hossain Mohd Rafie Johan Nor Qhairul Izzreen Mohd Noor |
| author_facet | Mohd Hazim Mohd Yusop Mohd Fadzelly Abu Bakar Kamarul Rahim Kamarudin Nur Fadhilah Khairil Mokhtar Mohd Abd Motalib Hossain Mohd Rafie Johan Nor Qhairul Izzreen Mohd Noor |
| author_sort | Mohd Hazim Mohd Yusop |
| collection | DOAJ |
| description | Point-of-care diagnostic methods for animal species determination are critical for rapid, simple, and accurate enforcement of food labelling. PCR is the most common method for species identification. However, the requirement of using a thermal cycler created drawbacks for the PCR application, particularly in low-resource settings. Hence, in this study, a method for porcine DNA detection using recombinase polymerase amplification (RPA), coupled with nucleic acid lateral flow immunoassay (NALFIA), was developed. Porcine-specific primers targeting pig (<i>Sus scrofa</i>) cytochrome b gene fragments specifically amplify a 197 bp fragment of the mitochondrial gene as being visualized by 2% agarose gel and PCRD NALFIA. The reaction temperature and time were 39 °C and 20 min, respectively. Herein, the specificity of the primers to porcine was confirmed after being assayed against six animal species, namely cow, goat, chicken, duck, dog, and rabbit. The porcine-specific RPA assay shows a high limit of detection of 0.01 ng/µL pork DNA. Based on the preliminary performance data obtained from this study, the potential of this method as a rapid and sensitive tool for porcine DNA detection in meat-based products is foreseen. |
| first_indexed | 2024-03-09T17:41:10Z |
| format | Article |
| id | doaj.art-f4f42c84e2a64c8e858d2916cfbc96a7 |
| institution | Directory Open Access Journal |
| issn | 1420-3049 |
| language | English |
| last_indexed | 2024-03-09T17:41:10Z |
| publishDate | 2022-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Molecules |
| spelling | doaj.art-f4f42c84e2a64c8e858d2916cfbc96a72023-11-24T11:36:58ZengMDPI AGMolecules1420-30492022-11-012723812210.3390/molecules27238122Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal AuthenticationMohd Hazim Mohd Yusop0Mohd Fadzelly Abu Bakar1Kamarul Rahim Kamarudin2Nur Fadhilah Khairil Mokhtar3Mohd Abd Motalib Hossain4Mohd Rafie Johan5Nor Qhairul Izzreen Mohd Noor6Faculty of Applied Science and Technology, Universiti Tun Hussein Onn Malaysia, Hub Pendidikan Tinggi Pagoh, Muar 84600, MalaysiaFaculty of Applied Science and Technology, Universiti Tun Hussein Onn Malaysia, Hub Pendidikan Tinggi Pagoh, Muar 84600, MalaysiaFaculty of Applied Science and Technology, Universiti Tun Hussein Onn Malaysia, Hub Pendidikan Tinggi Pagoh, Muar 84600, MalaysiaHalal Products Research Institute, Universiti Putra Malaysia, Putra Infoport, Serdang 43400, MalaysiaNanotechnology and Catalysis Research Centre, Institute for Advanced Studies, Universiti Malaya, Kuala Lumpur 50603, MalaysiaNanotechnology and Catalysis Research Centre, Institute for Advanced Studies, Universiti Malaya, Kuala Lumpur 50603, MalaysiaFaculty of Food Science and Nutrition, Universiti Malaysia Sabah, Jalan UMS, Kota Kinabalu 88400, MalaysiaPoint-of-care diagnostic methods for animal species determination are critical for rapid, simple, and accurate enforcement of food labelling. PCR is the most common method for species identification. However, the requirement of using a thermal cycler created drawbacks for the PCR application, particularly in low-resource settings. Hence, in this study, a method for porcine DNA detection using recombinase polymerase amplification (RPA), coupled with nucleic acid lateral flow immunoassay (NALFIA), was developed. Porcine-specific primers targeting pig (<i>Sus scrofa</i>) cytochrome b gene fragments specifically amplify a 197 bp fragment of the mitochondrial gene as being visualized by 2% agarose gel and PCRD NALFIA. The reaction temperature and time were 39 °C and 20 min, respectively. Herein, the specificity of the primers to porcine was confirmed after being assayed against six animal species, namely cow, goat, chicken, duck, dog, and rabbit. The porcine-specific RPA assay shows a high limit of detection of 0.01 ng/µL pork DNA. Based on the preliminary performance data obtained from this study, the potential of this method as a rapid and sensitive tool for porcine DNA detection in meat-based products is foreseen.https://www.mdpi.com/1420-3049/27/23/8122porcine DNARPANALFIAmeatballhalal authentication |
| spellingShingle | Mohd Hazim Mohd Yusop Mohd Fadzelly Abu Bakar Kamarul Rahim Kamarudin Nur Fadhilah Khairil Mokhtar Mohd Abd Motalib Hossain Mohd Rafie Johan Nor Qhairul Izzreen Mohd Noor Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal Authentication Molecules porcine DNA RPA NALFIA meatball halal authentication |
| title | Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal Authentication |
| title_full | Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal Authentication |
| title_fullStr | Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal Authentication |
| title_full_unstemmed | Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal Authentication |
| title_short | Rapid Detection of Porcine DNA in Meatball Using Recombinase Polymerase Amplification Couple with Lateral Flow Immunoassay for Halal Authentication |
| title_sort | rapid detection of porcine dna in meatball using recombinase polymerase amplification couple with lateral flow immunoassay for halal authentication |
| topic | porcine DNA RPA NALFIA meatball halal authentication |
| url | https://www.mdpi.com/1420-3049/27/23/8122 |
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