| Bovine spongiform encephalopathy (BSE) and scrapie have do harm to the health of human being and animals and cause huge damage to the global stockbreeding. The epidemiologic survey has proved that this disease caused by prions can be transmitted through food chain and is related to Creutzfeldt-Jakob Disease. The theory of feed-borne contamination through infected ruminant protein being the major vehicle for the transmission of BSE in cattle has been widely supported by studies, so many countries including China enforce the ban on meat-and-bone for farmed animals and it is necessary to detect the ruminant proteins in feedstuffs.Now there are four different analytical methods currently applied in the world for the detection and identification of animal tissues in feed including our country, i.e. microscopic analysis, polymerase chain reaction, immunoassay analysis and near infrared spectroscopy. There still have some disadvantages in these methods, e.g. the small number of samples, false positive results and much experience of operators. The tool of Real-time Fluorescence Quantitive PCR is a much better analytical method which is more quick, accurate and economical than them.This paper has three major research contents: the first is the conditions of extraction and purification of DNA of ruminant proteins in feedstuffs;the second is the reaction conditions of Real-time Fluorescence Quantitive PCR;the last is the specificity, sensitivity and stability of this method. The authors successfully establish a new approach through solving some keys as follow: the time, temperature and reagent of extraction of DNA; the sequence of primer and probe; the time and temperature of denaturation, annealing and elongation; the number of cycles.Real-time Fluorescence Quantitive PCR not only has no cross-contamination problem of classical PCR but also has good specificity, sensitivity and stability. It provides supporting data for feed safety supervision. |
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