The invention relates to a biological detection
reagent capable of discriminating three types of
ruminant animal varieties simultaneously, a preparation method and an application thereof. The invention selects the specificity of the cow, goat and sheep, and the conservative sequence segment of conservative mitochondrial
gene as target, applies Primer Eexpress 3.0
software and Primer Select
software in DNAStar, and designs and synthesizes a plurality of groups of primers and probes. After being synthesized and marked, the plurality of pairs of designed primers and the probes are carried out best
pairing screening experiment so as to obtain the fittest combination of primers and probes. The
reagent contains two pairs of specificity primers, and three Taq Man probes, wherein one pair of theprimer aims at cow source components, the other pair of primer aims at common primers of goats and sheep source component detection; the three probes are respective source components to the cows, thegoats and the sheep; and the amplified target fragment length to the source component detection of the cows, the goats and the sheep is respectively 92bp, 136bp and 136bp. The triple Taq Man
fluorescence PCR detection method can simultaneously carry out accurate quantitative detection to the source components of the cows, the goats and the sheep, has simple and convenient operation, intuitive result, strong specificity, high sensitivity and good
repeatability and can realize high
throughput detection of various source components.