Rapid detection kit and detection method for multiple virulence factor GeXP of vibrio coralliilyticus

A Coral-lysing Vibrio and detection kit technology, applied in the biological field, can solve the problems of cumbersome process, easy to miss detection, unable to fully reflect the potential pathogenicity of pathogenic bacteria, etc.

Active Publication Date: 2013-01-30
SOUTH CHINA SEA INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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Problems solved by technology

However, these methods have certain defects: (1) The detection is time-consuming and the process is cumbersome; (2) The detection sensitivity is low, and the pathogenic bacteria are often detected in the late stage of disease; (3) The target gene for detection is too single, and it is easy to miss detection. Different strains of the same species caused by environmental differences cannot fully reflect the potential pathogenicity of the pathogenic bacteria

Method used

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  • Rapid detection kit and detection method for multiple virulence factor GeXP of vibrio coralliilyticus
  • Rapid detection kit and detection method for multiple virulence factor GeXP of vibrio coralliilyticus
  • Rapid detection kit and detection method for multiple virulence factor GeXP of vibrio coralliilyticus

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Embodiment 1

[0077] (1) Genomic DNA extraction of Vibrio corallilyticus

[0078] Firstly, the Vibrio coralliilyticus in the coral sample was purified and isolated on a plate, and then the Vibrio coralliilyticus was cultured.

[0079] ① Add 50 μL of 10% (w / v) sterile Chelex-100 solution into the centrifuge tube;

[0080] ② Pick a single colony from the culture plate of Vibrio coralliilyticus and put it into the above Chelex-100 solution;

[0081] ③ Place the centrifuge tube on a vortex mixer and shake it fully for 10 seconds, take a boiling water bath for 10 minutes, cool to room temperature, and shake it fully for another 10 seconds;

[0082] ④ Centrifuge at 13000r / min for 2min, and the supernatant is the genomic DNA of Vibrio corallilyticus, and store it in a -20°C refrigerator for later use.

[0083] (2) Design and synthesis of multigene PCR primers

[0084] Primers were designed for the virulence-related gene flhA, and the amplified fragment size was about 147bp:

[0085] flhA-F: AG...

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Abstract

The invention discloses a rapid detection kit and a detection method for multiple virulence factor GeXP of vibrio coralliilyticus. Directed at 20 known vibrio coralliilyticus virulence genes, specific primers are designed. 20 genes can be detected for one time rapidly, efficiently and simultaneously by using the detection kit, according to the detection method and through the single-tube reaction, and accordingly, whether samples or environments contain the vibrio coralliilyticus carrying virulence relevant genes or having potential pathogenic capacities is detected. According to the rapid detection kit and the detection method for multiple virulence factor GeXP of vibrio coralliilyticus, the detection coverage is wide, the accuracy of vibrio coralliilyticus pathogenicity evaluation can be improved greatly, an effective detecting method is provided for vibrio coralliilyticus pathogenicity evaluation, and a large significance is provided for monitoring and preventing and treating coral bleaching diseases.

Description

Technical field: [0001] The invention relates to the field of biotechnology, in particular to a rapid detection kit and detection method specially used for the pathogenic Vibrio-Vibrio corallilyticus (Vibrio corallilyticus), which can cause coral bleaching. Background technique: [0002] Vibrio coralliilyticus is a halophilic gram-negative bacterium belonging to the genus Vibrio in the family Vibrio. It is a rod-shaped bacterium that cannot produce spores. It is motile because of a single flagella. It colonizes on marine agar It is cream-colored and yellow on TCBS medium. The edges of the colonies are neat and smooth. The optimum growth temperature is 30 degrees Celsius. It was first isolated from the bleached coral Pocillopora damicornis in the Indian Ocean, and was later isolated from the bleached corals in the Pacific Ocean and the Red Sea. isolated and also found in juvenile North Atlantic oysters. [0003] In the past 30 years, frequent outbreaks of various coral disea...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12R1/63
Inventor 陈偿刘助红高磊任春华胡超群
Owner SOUTH CHINA SEA INST OF OCEANOLOGY - CHINESE ACAD OF SCI
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