TaqMan probe real-time fluorescence PCR (Polymerase Chain Reaction) detection of angiostrongylus cantonensis infected snails

Abstract

The invention discloses a TaqMan probe for real-time fluorescence PCR (Polymerase Chain Reaction) detection of angiostrongylus cantonensis infected snails on the basis of a modern molecular biological technology, which is a detection method applied to snails infected with angiostrongylus cantonensis. The angiostrongylus cantonensis is food-borne zoonosis and is infected mainly through eating infected snails by mistake. The invention aims to provide a rapid and accurate method for detecting the snails infected with the angiostrongylus cantonensis. The TaqMan probe is technically characterized in that (1) a primer(upstream) having the sequence of 5'-TGA TGC TTG TGC GGT TTT C-3', a primer- (downstream) having the sequence of 5'-CCGAAG GCA AAG CAC ACA C-3' and a probe having the sequence of 5'-CGC TAT ACA CGC ACA TTT GCC GG-3' are provided; (2) the total volume of a reaction system is 20 mu L, wherein the reaction system comprises 10 muL of 2*Taq PCR MasterMix, 1 muL of Taqman probe, 1 muL of the upstream primer, 1 muL of the downstream primer, 1 mu L of template and the balance of ddH2O; and (3) in a reaction process, the pre-degeneration time is 3min at 94 DEG C, 10s at 94 DEG C and 30s at 60 DEG C, and the circulation is carried out for 40 times. The invention provides a new technological measure for food quarantine as well as investigation and monitoring of epidemic situations and has far reaching importance on preventing and controlling the occurrence of the angiostrongylus cantonensis diseases.

Description

1 technical field [0001] The invention relates to a novel molecular biology detection method, specifically, a real-time fluorescent PCR detection method for snails infected with Angiostrongylus cantonensis (Angiostrongylus cantonensis, AC). 2 background technology [0002] Angiostrongylus cantonensis is a new food-borne zoonotic disease. Frequent outbreaks of the disease were caused by people mistakenly eating shellfish such as apple snails, field snails, ring-shaped snails, and brown cloud agate snails. Up to now, there is still no report on the detection of Angiostrongylus cantonensis from intermediate hosts by real-time fluorescent PCR method. Traditional detection methods are still stuck in the stages of manual separation, microscopic examination, and expert identification. Not only the detection cycle is long, but also the shortcomings of extremely low sensitivity cannot be widely used. The applicant has successfully established a conventional PCR method to detect sna...

AI Technical Summary

Problems solved by technology

Traditional detection methods are still stuck in the stages of manual separation, microscopic examination, expert identification, etc. Not only the detection cycle is long, but also the shortcomings of extremely low sensitivity cannot be widely used

The applicant has successfully established a conventional PCR method to detect snails infected with Angiostrongylus cantonensis, which has improved the detection cycle and sensitivity, but it still cannot meet the emergency treatment of disease-causing emergencies, nor can it meet the requirements of shellfish food quarantine and epidemic outbreaks Rapid and accurate detection needs in investigation and surveillance

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