Primer group utilizing LAMP to detect vibrio alginolyticus and rapid diagnosis kit employing primer group

A technology for rapid diagnosis of Vibrio alginolyticus, applied in the direction of recombinant DNA technology, microbial measurement/inspection, and resistance to vector-borne diseases, etc., can solve the problems of lack of effective measures for prevention, cumbersome experimental operations, and affecting the detection time limit, etc. Achieve the effect of low detection cost, simple experimental device and strong operability

Inactive Publication Date: 2013-01-02
哈尔滨海关技术中心
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Problems solved by technology

[0003] At present, my country lacks effective measures for the prevention of bacterial pathogenic bacteria such as Vibrio alginolyticus in aquaculture, mainly relying on the widespread use of antibiotics, which has led to the emergence of drug-resistant strains and also poses a threat to human health
In addition, the detection methods

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  • Primer group utilizing LAMP to detect vibrio alginolyticus and rapid diagnosis kit employing primer group
  • Primer group utilizing LAMP to detect vibrio alginolyticus and rapid diagnosis kit employing primer group
  • Primer group utilizing LAMP to detect vibrio alginolyticus and rapid diagnosis kit employing primer group

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Embodiment Construction

[0032] The present invention will be further described below in conjunction with specific embodiment:

[0033] 1. Preparation of LAMP template

[0034] Extract and purify the bacterial genomic DNA according to the instructions of the TIANamp Bacteria DNA Kit, and add the corresponding reagents in proportion:

[0035] 1.1 Take 1.5mL of the bacterial enrichment solution to be tested, centrifuge at 10000r / min for 1min, discard the supernatant, add 200μL buffer GA, and suspend the bacteria thoroughly;

[0036] 1.2 Add 20 μL proteinase K (20mg / mL) and 220 μL buffer GB, mix thoroughly, and act in a 70°C water bath for 10 minutes;

[0037] 1.3 Add 220 μL of absolute ethanol, mix well for 15 seconds, centrifuge for 5-10 seconds, transfer the obtained solution (including flocculent precipitate) to the adsorption column CB3, centrifuge at 12000r / min for 30 seconds, and discard the liquid in the collection tube;

[0038] 1.4 Add 500μL buffer GD (containing absolute ethanol) to the adso...

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Abstract

The invention relates to a primer group utilizing LAMP to detect vibrio alginolyticus and a rapid diagnosis kit employing the primer group. The primer group comprises F3: CCAGTGGCTTACTCGTTGG, B3: TTCGCGGCCATAAACTCAT, FIP; CGTTCCACTGCCCACCAAACAAGCCCAGCACTGGTATATGC, and BIP: GGCAACCAAACGGACCTTGCCCGATTGATGACGCCCTTAG, and the rapid diagnosis kit comprises the primer group, positive reference substance, LAMP reaction liquid and BstDNAPolymerase polymerase. The primer group and the rapid diagnosis kit have the advantages that the operability is high, the operation speed is high, the result is visual to judge, the detection cost is low, and the experimental device is simple, and only a common water bath kettle or other equipment capable of obtaining stable heat sources is required.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to a primer set for detecting Vibrio alginolyticus by using LAMP and a rapid diagnostic kit thereof. Background technique [0002] Vibrio alginolyticus ( Vibrio alginolyticus ) is a halophilic Gram-negative bacterium that is widely distributed in estuaries and marine environments. Symptoms such as infection, gastroenteritis and sepsis seriously threaten the development of the aquaculture industry and cause huge economic losses. In recent years, due to the continuous occurrence of food poisoning incidents caused by Vibrio alginolyticus, people have further realized the pathogenicity and harm of Vibrio alginolyticus to humans, and therefore included it in the category of foodborne pathogens, becoming Food hygiene daily monitoring and quarantine objects. [0003] At present, my country lacks effective measures to prevent bacterial pathogens such as Vibrio al...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12N15/11
CPCY02A50/30
Inventor 徐义刚吴岩李丹丹刘忠梅刘新亮李苏龙
Owner 哈尔滨海关技术中心
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