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Method for quickly detecting pathogen of skin ulcerative syndrome in Apostichopus japonicus on site and kit thereof

A syndrome and kit technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., to achieve the effect of accurate detection, easy operation, and prevention of the spread of pathogens

Inactive Publication Date: 2014-09-24
大连市水产技术推广总站
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the detection of non-sluggish Edwardsiella pathogens did not produce specific amplification, thus proving that the detection of LAMP method has strong specificity
So far, there is no relevant report on the application of LAMP method to the detection of the pathogen (Vibrio splendidus) of sea cucumber rotting skin syndrome

Method used

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  • Method for quickly detecting pathogen of skin ulcerative syndrome in Apostichopus japonicus on site and kit thereof
  • Method for quickly detecting pathogen of skin ulcerative syndrome in Apostichopus japonicus on site and kit thereof

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Embodiment Construction

[0030] The kit of the embodiment of the present invention has a kit outer packaging (78×78×50) outside the kit, and there are in the kit:

[0031] 1. Five sample collection tubes numbered I, II, III, IV, V respectively;

[0032] 2.5 grinding rods;

[0033] 3.25 toothpicks;

[0034] 4. 7 sample sampling tubes numbered I, II, III, IV, V, (+) (positive control), (-) (negative control) respectively, containing FTA card membranes cut into small pieces;

[0035] 5.1 straws;

[0036] 6.1 The test solution tube filled with absolute ethanol;

[0037] 7. The numbers are I, II, III, IV, V, (+) (positive control), (-) (negative control) 7 membrane rinse tubes with high-purity water inside;

[0038] 8. The numbers are respectively I, II, III, IV, V, (+) (positive control), (-) (negative control) 7 bottles with TE buffer (containing 10mM Tris-HCl and 1mM EDTA, pH8.0) a nucleic acid denaturation tube;

[0039] 9. The numbers are respectively I, II, III, IV, V, (+) (positive control), (...

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Abstract

The invention discloses a method for quickly detecting pathogen of skin ulcerative syndrome in Apostichopus on site and a kit thereof. In the method, two pairs of primers, particular operation steps and the kit aiming at the vibrio splendidus are designed by using a loop-mediated isothermal amplification (LAMP) method. The DNA sequences of the reaction primer are as follows: VS-F3 is 5'-TGGTCCCCCCATATTCAGAC; VS-B3 is 5'-AAGTACCCCGAGGAAGAGAA; VS-FIP is 5'-GCCCCGTAACCGACACATCATTTTAGGATATCACGTGTCCCGC; and VS-BIP is 5'-GGCACTTTCCAGAGCCTTCGTTTTTAGATTCCGAAAGTAGCGGC. By using the method, expensive instrument and equipment or long detecting time is not required, the pathogen can be detected even less pathogen exists. The method has the advantages of low cost, accurate detection and the like, is convenient to operate, real-time monitor can be performed, and the epidemic situation can be discovered in time, the incidence trend of the epidemic disease is reduced, the pathogen can be prevented from distributing in early stage, and more loss brought to raisers is avoided.

Description

Technical field: [0001] The present invention relates to a kind of LAMP detection method of sea cucumber rotting skin syndrome pathogen (Vibrio resplendent), especially a kind of simple and convenient operation, low cost, accurate detection, can detect the method of sea cucumber rotting skin syndrome pathogen rapidly on the spot and Reagent test kit. Background technique: [0002] Sea cucumber rotting skin syndrome is an explosive epidemic, with a high infection rate and a mortality rate of more than 90%, which has been endangering sea cucumber cultivation. Studies have shown that Vibrio splendidus is the main pathogen of sea cucumber rotting skin syndrome, and early detection of Vibrio splendidus is a prerequisite for effective prevention and treatment of sea cucumber rotting skin syndrome. At present, there are immunofluorescence technique (FAT), enzyme-linked immunosorbent assay (ELISA), molecular detection techniques (PCR, RFLP, AFLP, RAPD) etc. for the detection of Vib...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12Q1/04
Inventor 陈文博王洪军李勃丁勇刘彤郝佳宋晓阳
Owner 大连市水产技术推广总站
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