基于TaqMan实时荧光聚合酶链式反应和DNA条形码技术鉴别松茸及其伪品

Identification of Tricholoma matsutake and its counterfeits based on TaqMan real-time fluorescence polymerase chain reaction technology and DNA barcoding technology

中文关键词:  松茸  TaqMan实时荧光PCR  DNA条形码技术  快速检测  真实性鉴别

英文关键词:Tricholoma matsutake  TaqMan real-time fluorescence polymerase chain reaction  DNA barcoding technology  rapid detection  authenticity identification

中文摘要:

      目的 建立松茸(Tricholoma matsutake)成分的TaqMan实时荧光聚合酶链反应(polymerase chain reaction, PCR)定性检测方法, 并结合DNA条形码技术鉴定松茸及其制品成分与其标签的符合性。方法 以松茸的pol基因为靶基因, 设计特异性引物及探针, 对其特异性、灵敏度、重复性开展研究分析, 并将建立的松茸成分TaqMan实时荧光PCR检测方法对不同种类市售松茸样品开展检测, 鉴别其真实性。结果 采用建立的TaqMan实时荧光PCR方法进行检测, 松茸与其他32种食用菌和动植物样品均无交叉反应, 方法特异性强; 灵敏度为质量分数0.01%的松茸和0.01 ng/μL松茸基因组DNA; 对市售160份松茸及其制品开展真实性检测, 研究表明松茸干品存在假冒现象, 检测结果与标签不符合率为36.70%, 采用DNA条形码技术进一步对假冒松茸的物种进行鉴定, 物种属性为皱环球盖菇(Stropharia rugosoannulata)、巨大口蘑(Tricholoma giganteum)和栎松口蘑(Tricholoma bakamatsutake), 其他收集的松茸制品成分检测结果中与标签不符合率分别为23.70%~60.00%。结论 该方法灵敏度高, 特异性强, 可快速精准地对松茸成分进行真实性和符合性鉴别。

英文摘要:

      Objective To establish a method for qualitative determination of Tricholoma matsutake ingredients by a TaqMan real-time fluorescence polymerase chain reaction (PCR), and identify the conformity of Tricholoma matsutake and its products with their labels by DNA barcoding technology. Methods Taking the pol gene of Tricholoma matsutake as the target gene, specific primers and probes were designed to study and analyze their specificity, sensitivity and repeatability. The established TaqMan real-time PCR method of Tricholoma matsutake was used to detect different types of commercially available samples and identify their authenticity. Results The established method was used for real-time PCR detection, and there was no cross-reactivity between Tricholoma matsutake and 32 kinds of other edible fungi, animals and plants, indicading strong specificity of the method. The sensitivity was 0.01% Tricholoma matsutake and 0.01 ng/μL Tricholoma matsutake genomic DNA. The authenticity test of 160 commercially available Tricholoma matsutake and their products showed that there was counterfeiting of dried Tricholoma matsutake. The test results did not match the labels at a rate of 36.70%. DNA barcoding technology was used to further identify the species of counterfeit matsutake mushrooms, with species attributes of Stropharia rugosoannulata, Tricholoma giganteum and Tricholoma bakamatsutake. The non-compliance rates with labels in the authenticity testing resultes of other collected matsutake products were 23.70%–60.00%, respectively. Conclusion This method has high sensitivity and strong specificity, which can quickly and accurately identify the authenticity and compliance of Tricholoma matsutake ingredients.