摘要
目的建立Taqman实时荧光PCR法对奶制品中掺加物大米、玉米、小麦、高粱、大麦等谷物源性成分的初筛检测体系。方法以常见掺加物大米、玉米、小麦、高粱、大麦等谷物的叶绿体rbcL基因作为靶基因,通过BLAST软件比对,选择其同源保守区域设计引物和探针。经通用性、特异性、灵敏性试验对探针和引物可行性进行验证,并通过模拟含谷物奶粉及市售奶类制品检测对其实际检测能力进行验证。结果所建立体系可扩增大米、玉米、小麦、高粱、大麦等常见的谷物掺加物的DNA提取物,与市售奶制品的主成分牛奶、羊奶以及其常见添加物香蕉、红枣、菠萝、草莓、西红柿、花生、大豆等动植物源性成分无交叉扩增(Ct>35)。对小麦纯DNA提取物检出限为0.01 ng。对分别掺加5种谷物成分的模拟奶制品检出限均可达0.5%,对市售的14份奶类食品中的谷类成分的检测结果均与食品标签相符。结论本研究建立的Taqman实时荧光PCR体系具有通用、相对特异、灵敏、实用等优点,可用于奶制品中掺加或掺杂掺假谷物源性成分的快速定性检测。
Objective A Taqman real-time PCR was developed to detect cereal-derived ingredients adulterated in dairy products. Methods The universal primers and probe for cereals were designed according to the homogeneous region of rbcL gene by blasting the rbcL gene of rice, maize, wheat, sorghum and barley. And the assay was evaluated by universality and sensitivity test. Meanwhile, its practicability was verified using simulated samples and market samples. Results implicated that the primers-probe system could detect DNA fragments of rice, maize, wheat, sorghum and barley with no cross-reaction to banana, jujube, pineapple, strawberry, tomato, peanut, soybean, bovine, ovine which may occur in dairy products （ Ct 〉35）. The detection limit was 0. 01 ng for pure wheat DNA and 0.5% for each of five cereals in dairy mixtures. 14 market dairy samples were analyzed for cereal ingredients, and were all consistent with their food labels. Conclusion The study suggested that the developed Taqman real-time PCR method was a rapid, sensitive and efficaciousdetection assay for cereal-derived ingredients in dairy products.
出处
《中国食品卫生杂志》
北大核心
2014年第2期123-127,共5页
Chinese Journal of Food Hygiene
基金
苏州市科技支撑计划项目(SS201126)
