Detection reagent, kit as well as application of detection reagent and kit

Abstract

The invention discloses a detection reagent, a kit as well as application of the detection reagent and the kit. The detection reagent comprises any one of or a combination of more of the following four primer pairs: a first primer pair comprising a first primer and a second primer with sequences shown in SEQ ID NO: 1 and SEQ ID NO:2 respectively, a second primer pair comprising a third primer anda fourth primer with sequences shown in SEQ ID NO:3 and SEQ ID NO:4 respectively, a third primer pair comprising a fifth primer and a sixth primer with sequences shown in SEQ ID NO:5 and SEQ ID NO:6 respectively as well as a fourth primer pair comprising a seventh primer and an eighth primer with sequences shown in SEQ ID NO:7 and SEQ ID NO:8 respectively. The detection method amplifies gene fragments in proper length by marked specific primers, can guarantee high specificity of detection and improve accuracy of detection results.

Description

technical field [0001] The invention relates to a detection reagent and a kit, in particular to a detection reagent, a kit and an application thereof for detecting E. The technical field of immunology. Background technique [0002] Escherichia coli, Staphylococcus aureus, Streptococcus dysgalactiae, Streptococcus agalactiae and other pathogenic bacteria widely exist in nature, which can pollute food, water, etc., and then pose a threat to the life safety of humans and animals. [0003] In order to prevent the harm of such pathogenic bacteria, people have developed a variety of pathogenic bacteria detection techniques. For example, for Escherichia coli, Staphylococcus aureus, Streptococcus dysgalactiae, and Streptococcus agalactiae, the current conventional detection method includes steps such as sample processing, enrichment culture, isolation culture, and staining observation. This detection method is the "gold standard" of clinical detection. ", but the workload is heavy...

AI Technical Summary

Problems solved by technology

For example, for Escherichia coli, Staphylococcus aureus, Streptococcus dysgalactiae, and Streptococcus agalactiae, the current conventional detection method includes steps such as sample processing, enrichment culture, isolation culture, and staining observation. This detection method is the "gold standard" of clinical detection. ", but the workload is heavy, and the detection cycle takes about a week

[0004] Others such as physiological and biochemical identification are relatively accurate, but the disadvantage is that the operation is complicated and time-consuming, but it is also time-consuming

Chromogenic plate detection can effectively detect E. coli, but the disadvantage is that it is time-consuming, requires professional operation, and the reagents are expensive

Membrane filtration technology is a quantitative test and has many of the same advantages as test tube fermentation technology. The most obvious advantage of membrane filtration technology over test tube fermentation technology is that it is very convenient to detect large volumes of water samples, which can increase the detection sensitivity and Reliability, the disadvantage is that the specificity is not high, and the results are easily affected by other bacteria in the water sample and misjudged. Generally, it is used as a speculative experiment and needs to be further confirmed by other methods; The traditional method saves time and labor (18-24h), and has high specificity. The disadvantage is that the cost of reagents is high; the biggest problem with immunological methods is the cross-reactivity of commercial monoclonal or polyclonal antibodies with various intestinal bacteria. It is easy to cause false positives, and it is necessary to prepare more specific monoclonal or polyclonal antibodies

[0005] In this way, the method based on molecular biology has unique advantages. Molecular biology identification can overcome the above shortcomings. Pathogens can be detected within a few hours, and the operation is simple and convenient; There are more and more researches. On the one hand, compared with traditional microorganism isolation, cultivation and identification methods, PCR detection technology reduces the length of the detection cycle and saves the cost of detection; it is suitable for clinical detection of Escherichia coli. Epidemiological surveys and investigations on the infection of individual dairy cows and herds of cows in the dairy herd are an effective and good method for the early prevention and control of E. On the one hand, PCR detection technology has high specificity and sensitivity, but the disadvantage is that this method needs to combine electrophoresis tank, agarose gel and other instrument reagents, use toxic substances such as nucleic acid dyes, and later analysis needs to calculate the gel imager, which is very important for the experiment. High security requirements and toxic

[0006] However, the existing detection methods have disadvantages such as complicated operation, easy to be affected by the environment, easy to infect, unsuitable for early diagnosis, and large error in detection results.

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