A multiplex-conventional PCR assay for bovine,ovine, caprine and fish species identification in feedstuffs: Highly sensitive and specific |
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| Affiliation: | 1. Institute for Biological Instrumentation of the Russian Academy of Sciences, Pushchino, Moscow region 142290, Russia;2. Structural Bioinformatics Laboratory, Faculty of Science and Engineering, Åbo Akademi University, Turku 20520, Finland;3. Pharmaceutical Sciences Laboratory, Faculty of Science and Engineering, Pharmacy, Åbo Akademi University, Turku 20520, Finland;4. Department of Molecular Medicine, USF Health Byrd Alzheimer''s Research Institute, Morsani College of Medicine, University of South Florida, Tampa, FL 33612, USA |
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| Abstract: | A multiplex PCR assay was developed for rapid and reliable identification of bovine, ovine, caprine and fish species in feedstuffs simultaneously. The method merges the use of bovine, ovine, caprine and fish primers that amplify fragments (ovine; 119 bp, caprine; 142 bp, fish; 224 bp and bovine; 271 bp) of the mitochondrial t.glu gene forward and cyt b reverse, 12S rRNA, 12S rRNA, and ATPase subunit 8 genes respectively, and a universal 18S rRNA primers that amplifies a 99 bp from eukaryotic DNA. To evaluate the effect of heat treatment, a severe sterilization condition (133 °C at 300 kPa for 20 min) was applied. Multiplex analysis of the reference feedstuff samples showed that the detection limit of the assay was 0.01% for each species. Taken together, all data indicated that this multiplex PCR assay was a simple, rapid, sensitive, specific, and cost-effective detection method for bovine, ovine, caprine and fish species in feedstuffs. |
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| Keywords: | Multiplex analysis Species-specific PCR Mitochondrial DNA 18S rRNA Authenticity Feedstuffs |
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