鸭血制品中猪、鸡、鸭源性成分的多重实时荧光定量聚合酶链式反应检测方法建立与应用OA

To address the adulteration of animal-derived components in duck blood products,this study developed a TaqMan probe-based real-time fluorescence quantitative polymerase chain reaction(real-time PCR)assay for identifying pig(Sus scrofa),chicken(Gallus domesticus),and duck(Anas platyrhynchos)-derived components in duck blood curd.Universe primers were designed based on the conserved regions of mitochondrial 16S rDNA sequences from these species,as well as species-specific molecular beacon probes(Z-Probe,J-Probe and Y-Probe)for their variable regions.The reaction conditions were optimized and the PCR assay was validated for specificity and sensitivity,and established a real-time PCR detection system.Furthermore,it was used to detect the adulteration of 19 non-prepacked duck blood curd samples from local markets.This assay allowed simultaneous differentiation of species-specific fluorescence signals in a single-tube reaction,demonstrating high specificity and sensitivity with an absolute limit of detection(LOD)of 0.06 ng/μL and a relative LOD of 0.1%in duck blood products.Out of the 19 samples,one was positive for pig-derived components,11 were positive for chicken-derived components,and 12 were positive for duck-derived components.This result was consistent with that of the real-time PCR method specified in the Chinese National Standard(GB/T 38164-2019)Identification of animal ingredient from common livestock and poultry-Real-time PCR,but our method was more sensitive.In conclusion,this method can effectively and accurately identify porcine and chicken adulterants in duck blood products,providing an accurate and robust technical approach for multi-species identification of animal-derived products.