Development of a DNA-based screening method to detect cow milk in ewe,goat and buffalo milk and dairy products using PCR-LCR-EIA-technique

摘要

In this contribution, a DNA-based technique which combines PCR (Polymerase Chain Reaction), LCR (Ligase Chain Reaction) and EIA (Enzyme Immune Assay) is introduced to detect the presence of cow milk in ewe, goat and buffalo milk and corresponding cheese. It is based on the analysis of one differing nucleotide between cow species (position 8411 of the Bos taurus beta -casein gene) and sheep, goat and buffalo species. This technique has been developed using pure milk or various mixtures of cow with ewe, goat and buffalo milk. DNA, extracted from milk or cheese samples, serves as template to amplify an universal 13-casein PCR product. Subsequently a LCR with species specific primers is performed using the PCR product as a template. LCR-primers are labelled with biotin or digoxygenin for further sensitive detection by EIA. This screening-technique allows a clear discrimination of cow species from sheep, goat and buffalo species in milk and cheese. A first quantitative analysis for different mixtures indicates a sensitivity of 5% cow DNA in ewe, goat and buffalo DNA.