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Specific forward and reverse primers and probe for vomiting-causing bacillus cereus, detection kit and application of detection kit

A technology of Bacillus cereus and forward and reverse primers, applied in the field of detection, can solve the problems of high cost, cumbersome detection, and long cycle, and achieve the effect of high accuracy, short time consumption, and instant on-site detection

Pending Publication Date: 2020-11-27
JIANGSU OCEAN UNIV +1
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Problems solved by technology

[0003] The existing detection of food-borne emetogenic Bacillus cereus is usually through the culture method, which requires laboratory operations and a long cycle, and the detection is relatively cumbersome; detection by molecular means usually requires expensive equipment and professional expertise. Skilled experimenters have a higher cost; in recent years, due to the rise of recombinase polymerase isothermal amplification technology (RPA technology) and lateral flow chromatography test strip detection technology (LFS), the present invention quickly identifies emesis by RPA-LFS technology Bacillus cereus, in order to bring new technical references for the detection of this bacterium

Method used

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  • Specific forward and reverse primers and probe for vomiting-causing bacillus cereus, detection kit and application of detection kit
  • Specific forward and reverse primers and probe for vomiting-causing bacillus cereus, detection kit and application of detection kit
  • Specific forward and reverse primers and probe for vomiting-causing bacillus cereus, detection kit and application of detection kit

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Embodiment

[0028] experiment material

[0029] Non-emetic B.cereus, emetic B.cereus, B.thuringiensis, B.cytotoxicus, mushroom B. mycoides, B. pseudomycoides, B. amyloliquefaciens, B. subtilis, B. brevis, lichen Bacillus (B.licheniformis), Listeria monocytogenes (L.monocytogenes), Salmonella enteritidis (S.enteritidis), Staphylococcus aureus (S.aureus), Escherichia coli O157 (E.coli O157) standard strain genome kept by the laboratory; Basic kit, nfo kit was purchased from TwistDX Company in the UK; lateral flow chromatography test strips were purchased from Hangzhou Ustar Biotechnology Co., Ltd.; PCR cleaning kit was purchased from Mona Biotechnology Co., Ltd.; Qubit 4 was purchased from Thermo Scientific; primers used in this study Both probes and probes were synthesized by General Biosystems (Anhui) Co., Ltd.; Roche LightCycler480 fluorescent quantitative PCR instrument was purchased from Roche Pharmaceuticals.

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Abstract

The invention relates to the technical field of detection, in particular to specific forward and reverse primers and a probe for vomiting-causing bacillus cereus, a detection kit and application of the detection kit. The detection kit comprises two sets of forward and reverse primers, the probe and a lateral flow chromatography test strip; the 5' end of the reverse primer is marked by biotin; the5' end of the probe is marked by Fluorescein Isothiocyanate (FITC), the C basic group (at least 30bp away from the 5' end and at least 15bp away from the 3' end) in the middle area is replaced by [THF], and the 3' end is subjected to end sealing by C3-spacer. According to the invention, a method adopting RPA-LFS (The isothermal recombinase polymerase amplification and the lateral flow strip) can realize real-time, on-site and instant rapid field detection with high accuracy without depending on instrument settings in a non-laboratory environment.

Description

technical field [0001] The invention relates to the technical field of detection, in particular to an emetogenic bacillus cereus-specific forward and reverse primers and probes, a detection kit and applications thereof. Background technique [0002] Bacillus cereus (B.cereus) broadly refers to the genus Bacillus cereus, and narrowly refers to the specific species of Bacillus cereus, which is a Gram-positive, aerobic, rod-shaped, endophytic spores, and food-borne pathogens that are resistant to drying and high temperature. Bacillus cereus is widely distributed in various environments and foods due to its low nutritional requirements and high resistance to complex conditions. The spores of Bacillus cereus that survived food processing conditions such as drying, cooking, and pasteurization germinate rapidly at a suitable temperature of 10-60°C to produce toxins. There are two typical clinical symptoms caused by Bacillus cereus toxin: diarrhea and vomiting. Diarrhea is caused...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/689C12Q1/6844C12Q1/04C12N15/11G01N33/569G01N33/543C12R1/085
CPCC12Q1/689C12Q1/6844G01N33/54313G01N33/56911C12Q2521/507C12Q2522/101
Inventor 高嵩董井泉赵盼盼王蕾董宇杨潇含王昱
Owner JIANGSU OCEAN UNIV
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