This study aimed to compare the sensitivity of porcine-specific primers targeting different mitochondrial DNA for the detection of porcine gelatin in ramen stock powder designated “Halal” using SYBR Green I real-time PCR. Four mitochondrial DNA markers (D-loop, ATP8, ND5, and 12S rRNA) were used to detect the porcine gelatin DNA. Extraction of DNA from porcine gelatin and spiked mixtures was successfully performed using a DNeasy Mericon Food Kit. Each primer had high porcine specificity for D-loop, ATP8, and ND5 genes, but not the 12S rRNA gene. Also, the detection limit for each primer was tested in a serial dilution of porcine gelatin DNA (10, 1, 0.1, 0.05, 0.01, and 0.005 ng) and ramen stock powder spiked with porcine gelatin (1, 0.1, 0.05, and 0.025%). Assays targeting both D-loop and ND5 genes were able to detect a level as low as 0.01 ng, while the detection limit for the ATP8 gene was 0.05 ng in pure porcine gelatin. The detection limits of D-loop, ATP8, and ND5 genes were 0.05%, 0.1%, and 1% (w/w) in ramen stock powder spiked with porcine gelatin, respectively. In conclusion, a novel detection system targeting the mitochondrial D-loop was superior to the others and could be an effective and simple method to authenticate Halal products.

这项研究旨在比较针对不同线粒体DNA的猪特异性引物对使用SYBR Green I实时PCR检测名为“ Halal”的拉面贮藏粉中猪明胶的敏感性。四个线粒体DNA标记（D-loop，ATP8，ND5和12S rRNA）用于检测猪明胶DNA。使用DNeasy Mericon食品试剂盒成功地从猪明胶和加标混合物中提取了DNA。每个引物对D-loop，ATP8和ND5基因都具有很高的猪特异性，但对12S rRNA基因却没有。同样，在猪明胶DNA（10、1、0.1、0.05、0.01和0.005 ng）和掺有明胶（1、0.1、0.05和0.025的拉面原粉）的系列稀释液中测试每种引物的检出限％）。针对D-loop和ND5基因的检测方法能够检测到0.01 ng的水平，纯猪明胶中ATP8基因的检出限为0.05 ng。在掺有猪明胶的拉面储备粉中，D-loop，ATP8和ND5基因的检出限分别为0.05％，0.1％和1％（w / w）。总之，针对线粒体D环的新型检测系统优于其他检测系统，并且可能是鉴定清真产品的有效且简单的方法。