Primer probe group, kit and detection method for multiple detection of enteropathogenic bacteria based on fluorescent RMA method

A multiplex detection, primer-probe technology, applied in the field of molecular biology, can solve the problems of long detection time, high false positive rate and easy contamination of molecular biology detection methods

Pending Publication Date: 2021-05-07
济南国益生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Molecular biology detection methods have the disadvantages of lon

Method used

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  • Primer probe group, kit and detection method for multiple detection of enteropathogenic bacteria based on fluorescent RMA method
  • Primer probe group, kit and detection method for multiple detection of enteropathogenic bacteria based on fluorescent RMA method
  • Primer probe group, kit and detection method for multiple detection of enteropathogenic bacteria based on fluorescent RMA method

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Embodiment 1

[0050] 1. Preparation of positive standard plasmid

[0051]Extract the DNA of Salmonella, Shigella, and Vibrio parahaemolyticus, and use it as a template to perform PCR amplification on the specific genes of Salmonella, Shigella, and Vibrio parahaemolyticus, and the PCR amplification products are passed through 1% agarose gel Electrophoresis, gel tapping recovery, cloning and connection to pMD18-T vector, transformation into Escherichia coli competent cells, blue-white screening, picking white colonies, and colony PCR verification. Send the positive recombinant bacteria to the company for sequencing, culture the correctly sequenced recombinant bacteria overnight, extract the plasmid DNA, and obtain the positive plasmid.

[0052] 2. Design of fluorescent RMA primers and probes

[0053] Fluorescent RMA primers and probes were designed for the invA gene of Salmonella, the ipaH gene of Shigella, and the tlh gene of Vibrio parahaemolyticus, as shown in Table 1:

[0054] Table 1 P...

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Abstract

The invention belongs to the technical field of molecular biology, and particularly relates to a primer probe group, kit and detection method for multiple detection of enteropathogenic bacteria based on a fluorescent RMA method. The primer probe group comprises a primer and a probe for detecting salmonella, a primer and a probe for detecting shigella, and a primer and a probe for detecting vibrio parahaemolyticus.

Description

technical field [0001] The application belongs to the technical field of molecular biology, and specifically relates to a primer-probe set, a kit and a detection method for multiple detection of intestinal pathogenic bacteria based on the fluorescent RMA method. Background technique [0002] Enteropathogenic bacteria refer to bacteria that can cause disease except the normal flora of the large intestine, mainly including Salmonella, Shigella, Vibrio parahaemolyticus, Vibrio cholerae ( Vibrio cholerae), Clostridium difficile (Clostridium difficile) and Escherichia coli (Escherichiacoli) etc. When pathogenic bacteria enter the body and reproduce in large numbers, it will cause a series of intestinal reactions in the body, which seriously threatens human health. At the same time, infectious diseases caused by intestinal pathogenic bacteria are the main factors that endanger public health safety. The rapid and correct detection of intestinal pathogenic bacteria in clinical prac...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/6844C12Q1/04C12Q1/10C12N15/11C12R1/42C12R1/01C12R1/63
CPCC12Q1/689C12Q1/6844C12Q2600/16
Inventor 刘建生陈大为陈龙张瑶
Owner 济南国益生物科技有限公司
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