Camp detection primer set and kit for Bacillus cereus enterotoxin gene

A Bacillus cereus, detection primer technology, applied in the direction of microbial determination/inspection, recombinant DNA technology, biochemical equipment and methods, etc. Realize fast and accurate detection, shorten detection time, and have the effect of strong specificity

Active Publication Date: 2022-08-09
SHENYANG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there is no report on the CAMP detection technology of Bacillus cereus enterotoxin genes entFM and bceT

Method used

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  • Camp detection primer set and kit for Bacillus cereus enterotoxin gene
  • Camp detection primer set and kit for Bacillus cereus enterotoxin gene
  • Camp detection primer set and kit for Bacillus cereus enterotoxin gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] A kit for detecting Bacillus cereus, comprising a CAMP detection primer set of Bacillus cereus enterotoxin gene, the primer set includes two sets of primers, respectively for the amplification of Bacillus cereus enterotoxin gene entFM region fragment Primers (entFM-NF; entFM-NR) and primers for the amplification of the bceT region fragment of the Bacillus cereus enterotoxin gene (bceT-NF; bceT-NR);

[0037] The kit includes several CAMP reaction tubes containing CAMP working solution. The CAMP working solution is prepared by putting the above primer set into DEPC treated water; the kit also includes a CAMP reaction tube containing the entFM primer set and a bceT primer set. CAMP reaction tube. The CAMP reaction tube containing the entFM primer set, in which the concentrations of entFM-NF ​​and entFM-NR were both 1.6 μM, and the working solution volume was 1 μL; the CAMP reaction tube containing the bceT primer set, in which the concentrations of bceT-NF and bceT-NR were...

Embodiment 2

[0052] The following reactions were performed using the kit in Example 1.

[0053] The amplification reaction of Bacillus cereus enterotoxin gene bceT was verified using Eva Green chromogenic solution in the kit.

[0054] The total reaction solution system is 25 μL, and the combination is as follows:

[0055] 20mM Tris-HCl (pH8.8); 10mM KCl; 10mM (NH 4 ) 2 SO 4 ; 6mM MgSO 4 ; 0.1% Triton X-100; 1M betaine; 1.4mM dNTP; 8U Bst DNA polymerase; 1μL Eva green; 1μL nucleic acid sample;

[0056] Primer: 1 μL of CAMP working solution containing bceT primer set;

[0057] Target: Bacillus cereus-bceT dsDNA / GenBank: D17312.1;

[0058] A no-target control group was also set up.

[0059] The reaction conditions are as follows:

[0060] 1) Add the nucleic acid sample, CAMP reaction solution and DEPC treated water to a CAMP reaction tube containing the bceT primer set to obtain an amplification reaction solution;

[0061] 2) Set the reaction temperature of Applied Biosystems QuantSt...

Embodiment 3

[0065] A kit for detecting Bacillus cereus, comprising a CAMP detection primer set of Bacillus cereus enterotoxin gene, the primer set includes two sets of primers, respectively for the amplification of Bacillus cereus enterotoxin gene entFM region fragment Primers (entFM-NF; entFM-NR) and primers for the amplification of the bceT region fragment of the Bacillus cereus enterotoxin gene (bceT-NF; bceT-NR);

[0066] The kit includes several CAMP reaction tubes containing CAMP working solution. The CAMP working solution is prepared by placing the above primers in DEPC treated water; the kit also includes a CAMP reaction tube containing the entFM primer set and a CAMP containing the bceT primer set. reaction tube. CAMP reaction tube containing entFM primer set, in which the concentrations of entFM-NF ​​and entFM-NR are both 2.0 μM, and the working solution volume is 2.0 μL; CAMP reaction tube containing bceT primer set, in which the concentrations of bceT-NF and bceT-NR are Both ...

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Abstract

The present invention relates to a CAMP detection primer set and kit for Bacillus cereus enterotoxin gene. The primer set includes two sets of primers, namely, primers for amplifying the entFM region fragment of the Bacillus cereus enterotoxin gene and primers for amplifying the bceT region fragment of the Bacillus cereus enterotoxin gene. The primer set provided by the invention has high sensitivity and strong specificity, and the kit prepared by the primer set can quickly and accurately detect whether the sample contains Bacillus cereus, and at the same time judge whether it contains the toxin-producing genes entFM and bceT. Moreover, because the primer set provided by the present invention has extremely high specificity, the time required for CAMP amplification is short, which further shortens the detection time and simplifies the operation.

Description

technical field [0001] The invention belongs to the technical field of bacterial gene detection, and particularly relates to a CAMP detection primer set and a kit for Bacillus cereus enterotoxin gene. Background technique [0002] Bacillus cereus is a non-capsulated, motile, spore-forming, facultative aerobic, food-borne opportunistic pathogen, widely distributed in air, soil, sewage and various raw and cooked in food and food raw materials. After eating food contaminated with Bacillus cereus, people generally experience vomiting or diarrhea, or both symptoms of poisoning within 8 to 16 hours. Vomiting-type poisoning is mainly caused by DON, while diarrhea-type poisoning is caused by a variety of enterotoxins. Bacillus cereus poses a threat to food safety and human health and safety. Therefore, it is necessary to understand the potential toxicity of Bacillus cereus in food, and to establish a rapid and accurate detection scheme to monitor it. [0003] At present, the dete...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/689C12Q1/6844C12Q1/04C12N15/11
CPCC12Q1/689C12Q1/6844C12Q2531/119C12Q2537/1376Y02A50/30
Inventor 陈旭包海燕李薇石波乔宇彭晴岳喜庆乌日娜武俊瑞
Owner SHENYANG AGRI UNIV
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