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首页> 外文期刊>Poultry Science >Evaluation of a TaqMan real-time PCR assay for detection of chicken, turkey, duck, and goose material in highly processed industrial feed samples.
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Evaluation of a TaqMan real-time PCR assay for detection of chicken, turkey, duck, and goose material in highly processed industrial feed samples.

机译：TaqMan实时PCR分析的评估，以检测高度加工的工业饲料样品中的鸡肉，火鸡，鸭肉和鹅肉。

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摘要

A TaqMan real-time PCR method based on nucleotide sequence variation in the D-loop and 12S rRNA mitochondrial genes has been developed for the specific detection of chicken, turkey, duck, and goose prohibited material in animal feeds. The assay uses 4 primer/probe sets targeting short species-specific mitochondrial sequences together with a positive amplification control based on the eukaryotic 18S rRNA gene. The applicability of the real-time PCR assay was assessed through analysis of a batch of industrial feed samples subjected to different rendering temperatures according to European legislation regulations. The chicken-specific real-time PCR system allows a highly sensitive qualitative detection of chicken-derived processed animal protein from different tissue-type origins, even in samples containing 0.1% target and subjected to heat treatments higher than 133 degrees C. On the other hand, turkey, goose, and duck real-time PCR systems also allowed detection of as low as 0.1% target material in binary mixtures (muscle/oat) manufactured using the minimum legal requirements for sterilization temperatures (133 degrees C). Quantification results, based on calibration standard curves, were very reproducible under the experimental conditions tested. However, the quantitative capability of the assay is limited by the existing variability in terms of composition and processing treatment of the feeds, which affect the amount and quality of amplifiable DNA.
机译：已开发出一种基于D环和12S rRNA线粒体基因核苷酸序列变异的TaqMan实时PCR方法，用于特异性检测动物饲料中的鸡肉，火鸡，鸭和鹅禁用物质。该测定法使用4个针对短物种特异性线粒体序列的引物/探针集以及基于真核18S rRNA基因的阳性扩增对照。根据欧洲法规，通过对一批经受不同提炼温度的工业饲料样品进行分析，从而评估了实时荧光定量PCR检测的适用性。鸡特有的实时PCR系统可对不同组织类型来源的鸡衍生的动物蛋白进行高灵敏度的定性检测，即使在含有0.1％靶标且经过133摄氏度以上热处理的样品中也是如此。手动，火鸡，鹅和鸭的实时PCR系统还允许检测使用最低法律要求的灭菌温度（133摄氏度）生产的二元混合物（肌肉/燕麦）中低至0.1％的目标物质。基于校准标准曲线的定量结果在测试的实验条件下非常可重现。但是，测定的定量能力受到饲料组成和加工处理方面现有变异性的限制，这些变异性影响可扩增DNA的数量和质量。

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