Molecular motor biosensor kit used in vibrio parahaemolyticus molecular typing

A biosensor, Vibrio hemolyticus technology, applied in the direction of microorganism-based methods, microorganisms, microorganism determination/inspection, etc., can solve the problems of complex and cumbersome procedures, and achieve high sensitivity, high-throughput detection, and strong specificity. Effect

Active Publication Date: 2013-02-20
PEOPLES REPUBLIC OF CHINA BEIJING ENTRY EXIT INSPECTION & QUARANTINE BUREAU +1
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Problems solved by technology

However, the above-mentioned methods are relatively cumbe...

Method used

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  • Molecular motor biosensor kit used in vibrio parahaemolyticus molecular typing
  • Molecular motor biosensor kit used in vibrio parahaemolyticus molecular typing
  • Molecular motor biosensor kit used in vibrio parahaemolyticus molecular typing

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Embodiment

[0042] (1) Culture of bacteria and extraction of DNA

[0043] Ten strains of V.parahaemolyticus were isolated from food. Inoculate 10 strains of V.parahaemolyticus into APW containing 3% NaCL to activate and multiply, and culture at 36°C±1°C for 24h. Then streak inoculated on TCBS agar, cultured at 36°C±1°C for 24h. Finally, suspicious colonies were picked and transferred to TSA agar with 3% NaCL, and cultured at 36°C±1°C for 24h for DNA extraction.

[0044] The cultured V. parahaemolyticus was extracted with a DNA extraction kit (spin column type) DP320, and the detailed operation steps were referred to the kit instruction manual. The liquid in the finally obtained centrifuge tube is the DNA extraction liquid of the bacteria, which is used as a template during PCR amplification and as a target detection substance of the molecular motor sensor. The DNA extraction solution was stored in a refrigerator at -20°C, and was thawed naturally at room temperature when used, and then...

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Abstract

The invention belongs to a kit used for carrying out rapid molecular typing determination upon vibrio parahaemolyticus pathogenicity. Specifically, vibrio parahaemolyticus species-specific genes tlh and toxR and virulence genes tdh and trh are detected by using F0F1-ATPase molecular motor biosensors Chro-tlh, Chro-tdh, Chro-trh, and Chro-toxR in the kit. On the one hand, with the detections upon species-specific genes tlh and toxR, a target strain can be identified. On the other hand, with the detections upon tdh and trh, the target strain pathogenicity can be determined. A method comprises that: F0F1-ATPase molecular motor biosensors in the kit are constructed; molecular typing is carried out by using the F0F1-ATPase molecular motor biosensors; and vibrio parahaemolyticus pathogenicity is determined according to a typing result. The kit has the advantages of high sensitivity, high speed, simple operation, and low cost.

Description

technical field [0001] The invention belongs to a kit for rapid molecular typing and judging of the pathogenicity of Vibrio parahaemolyticus, specifically using F in the kit 0 f 1 -ATPase molecular motor biosensors Chro-tlh, Chro-tdh, Chro-trh, Chro-toxR detect species-specific gene tlh, toxR and virulence gene tdh, trh of Vibrio parahaemolyticus (Vibrio parahaemolyticus), thereby Make a judgment on whether V.parahaemolyticus is pathogenic. Background technique [0002] V. parahaemolyticus is a common food-borne pathogen in coastal areas, which can cause typical gastroenteritis reactions such as diarrhea, intestinal cramps, nausea, vomiting, and fever in patients. In recent years, the scale of food poisoning caused by it and the scale of population exposure have shown an obvious upward trend. Therefore, the harmfulness of V.parahaemolyticus cannot be ignored. [0003] Although most species of V. parahaemolyticus do not cause disease in humans, there are still certain cha...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12R1/63
Inventor 张捷李兆杰王静张惠媛汪琦张昕顾德周尚士进陆琳王佩荣陈广全乐加昌
Owner PEOPLES REPUBLIC OF CHINA BEIJING ENTRY EXIT INSPECTION & QUARANTINE BUREAU
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