PCR (polymerase chain reaction) method with internal amplification control for detecting bacillus cereus in food and kit

Abstract

The invention provides a PCR (polymerase chain reaction) method with internal amplification control for detecting bacillus cereus in food and a kit. Compared with the conventional method, the method has the advantages of convenience in use, good reliability, short detection period, high sensitivity, high specificity, low cost and simple operation steps, is applicable to high-throughput operation and standard operation and has great significance for improvement of food safety. The method comprises steps as follows: (1), a pair of specific primers A / B and a pair of internal amplification control primers C / D specific to the bacillus cereus are designed by primer premier 6.0 software; (2), suspicious samples are collected for enrichment culture, and after DNA (deoxyribonucleic acid) is extracted, the two pairs of primers A / B and C / D, internal amplification control templates and detection sample DNA are subjected to amplification jointly; (5), a PCR pre-mixed solution of the kit is taken to prepare a 25 mu l reaction system for amplification, and a detection result is observed under an ultraviolet lamp after agarose gel electrophoresis.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a PCR method and a kit for detecting bacillus cereus in foods with an amplified internal standard. Background technique [0002] Bacillus cereus is a spore-forming Gram-positive bacillus that is widely distributed in nature and often exists in soil, dust, sewage, plants and many raw and cooked foods. At present, the bacteria has been isolated from a variety of food species, including meat, dairy products, vegetables, fish, potatoes, paste, soy sauce, pudding, fried rice and various desserts. Human consumption of food contaminated by Bacillus cereus may cause food poisoning. Symptoms usually manifest as symptoms of gastrointestinal infection such as nausea, vomiting, and diarrhea, and can also cause parenteral infection such as meningitis, pneumonia, endocarditis and Systemic infection etc. Every year in our country, there are many food poisoning incidents caused by eating food cont...

AI Technical Summary

Problems solved by technology

my country's national standard detection method for Bacillus cereus (GB 4789.14-2014) and the entry-exit inspection and quarantine industry standard (SN / T 0176-2013) both use the culture method, which requires enrichment culture, isolation and pure culture, and biochemical identification The experimental process is time-consuming and labor-intensive, the specificity is not strong, and the sensitivity to bacteria in the "viable but non-culturable state" (Viable But Non-Culturable, VBNC) that is still pathogenic is low, which may lead to false negative test results

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