Puffer fish ingredient fluorescence quantitative PCR detection reagent and preparation method and application thereof

Abstract

The invention discloses a puffer fish ingredient fluorescence quantitative PCR detection reagent and a preparation method and application thereof, in the technical scheme, relatively conservative cytochrome B gene fragments of different puffer fishes are selected to be the target sequences, after elaborately design and synthesis, a pair of specific primer and a specific fluorescent probe are contained, the length of the target segment is enlarged to 102bp, the sequences of the primer and the probe are respectively as follows: primer: Takifugu TF:5'-TCTTCACGAAACAGGCTCCAAC-3', Takifugu TR: 5'-GTGAAGCCCAGGAGGTCTTTGTA-3'; and probe: 5'-FAM-CGCAGACAAAATCCCATTCCACCCATA-TAMRA-3'. The invention is the fluorescence quantitative PCR detection reagent with high specificity, strong sensibility and rapid detection to the puffer fish ingredient mixed with minced fish products; and the method of the invention has reasonable design and accurate definiteness.

Description

technical field [0001] The invention relates to a fluorescent quantitative PCR detection reagent, in particular to a fluorescent quantitative PCR detection reagent for puffer fish components and a preparation method and application thereof. Background technique [0002] Puffer fish is tender, delicious, and nutritious. It enjoys the reputation of "the king of fish", is favored by gourmets, and is also very popular in Japan and South Korea. However, puffer fish viscera, ovaries, blood, fish skin, fish head and other parts are highly toxic. Improper processing or ingestion will often cause poisoning and even death. Because ordinary people do not have the relevant professional knowledge, sometimes they will be poisoned and die because they cannot identify puffer fish, especially for various products that have been processed into fish fillets and surimi, it is difficult to identify them by human senses; In recent years, some people intentionally or unintentionally processed puff...

AI Technical Summary

Problems solved by technology

Biological identification and near-infrared spectroscopy are time-consuming, the workload is very heavy, and the detection sensitivity is not high

Since the bone glue and other substances contained in the feed sample will cross-react with the antibody and interfere with the ELISA result, the detection sensitivity of the ELISA method is low, so it can only be used as a coarse screening method

PCR is used to detect components mixed with other species. It has good sensitivity and specificity, but it needs post-PCR treatment such as DNA electrophoresis, and exposure to toxic reagents ethidium bromide is prone to cross-contamination and cause false positive results.

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