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Specific target, primer and detection method for detecting Escherichia coli O157:H7, and application of specific target and primer

A technology of Escherichia coli and O157, applied in biochemical equipment and methods, microbiological determination/testing, DNA/RNA fragments, etc., can solve problems that are difficult to cover widely and poor gene specificity

Pending Publication Date: 2021-02-02
NANJING AGRICULTURAL UNIVERSITY
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Problems solved by technology

Some genes are less specific and often difficult to broadly cover different subtypes of E. coli O157:H7

Method used

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  • Specific target, primer and detection method for detecting Escherichia coli O157:H7, and application of specific target and primer
  • Specific target, primer and detection method for detecting Escherichia coli O157:H7, and application of specific target and primer
  • Specific target, primer and detection method for detecting Escherichia coli O157:H7, and application of specific target and primer

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Embodiment Construction

[0074] The present invention will be further described below in conjunction with the examples. The experimental methods that do not indicate specific conditions in the following examples generally follow the known means in the art, or according to the manufacturer's suggested conditions, and the bacterial strains involved in the examples all belong to the prior art. , those skilled in the art can easily obtain from public or commercial sources.

[0075] 1. Discovery of specific targets of enterohemorrhagic E. coli O157:H7

[0076] 1.1 Local BLAST installation and E.coli O157:H7 genome database construction

[0077] Software installation: Enter NCBI official website ( https: / / www.ncbi.nlm.nih.gov / ), click AllResources, download the .BLAST offline installation package (https: / / ftp.ncbi.nlm.nih.gov / blast / executables / blast+ / LATEST / ), and download the corresponding installation package (*.exe) according to the computer system.

[0078] Database construction: Based on the 83 com...

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Abstract

The invention provides a specific target, primer and detection method for detecting Escherichia coli O157:H7, and an application of the specific target and primer. Compared with a conventional detection target, the novel target provided by the invention has relatively good specificity, so that the generation of false-positive results in the detection is reduced; and an Eva Green third-generation saturated fluorescent dye is adopted, the dye has a relatively high fluorescence signal value, and therefore the detection sensitivity is improved. Meanwhile, even if the dye is used at a high concentration, the dye does not show the PCR inhibition effect, so that the resolution of a melting curve during detection is greatly improved, and therefore the detection accuracy is further improved; the detection method is convenient and fast, the observation is convenient, tedious electrophoresis operations are not needed, and the detection time is shortened within 12 h; and no antibody is involved inthe detection process, the detection cost is low, and therefore detection requirements of general laboratories can be met.

Description

technical field [0001] The invention belongs to the technical field of food safety detection, and specifically relates to a specific target point and primer for detecting Escherichia coli O157:H7, combining it with a saturated fluorescent dye, and adopting a double HRM-RT-PCR method to detect intestinal hemorrhage in food A method for the detection of Escherichia coli O157:H7. Background technique [0002] There are many types of Escherichia coli, Enterohemorrhagic Escherichia coli (EHEC) is a kind of pathogenic Escherichia coli, and the main pathogenic strain is Escherichia coli O157:H7 (Escherichia coli O157:H7, E. coli O157:H7). E. coli O157:H7 is a flagellated, non-spore-forming, motile Gram-negative bacillus. It is a food-borne pathogen that seriously threatens the health of the population and public health safety. People infected with this bacteria often suffer from severe abdominal cramps and bloody diarrhea, and nearly one in ten patients develops hemolysis Chroni...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/686C12Q1/10C12N15/11
CPCC12Q1/689C12Q1/686C12Q2600/16C12Q2537/143C12Q2563/107C12Q2527/107C12Q2521/107Y02A50/30
Inventor 别小妹孔梁宇陆兆新胡安妥
Owner NANJING AGRICULTURAL UNIVERSITY
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