Streptococcus faecalis dry powder lamp rapid detection kit and its application method

Abstract

The invention discloses a Streptococcus faecalis powdering LAMP rapid detection kit, and a rapid detection method based on the kit. The kit comprises a freeze-dried isothermal amplification agent, a re-dissolving solution, a positive contrast dry powder, a negative contrast substance, a colorimetric solution, a lysate and a sealing solution, and the freeze-dried isothermal amplification agent contains dNTPs, Bst DNA polymerase, an amplimer and a freeze-drying protection agent. The primer in the kit corresponds to eight segments of a Streptococcus faecalis specific gene sequence, the colorimetric solution is added after a reaction, and a negative and positive colorimetric difference is obvious. The kit has the advantages of strong specificity, high sensitivity, easiness in operation and simplicity in result judgment, and is suitable for the quality control and the safe and rapid detection of foods, beverages, water and other raw and auxiliary materials. The kit is convenient for normal temperature transportation, is simple and fast to use without special devices, reduces the cost, widens the application range of products, is suitable for onsite rapid detection and base remote region related detection, and has wide application prospect.

Description

technical field [0001] The invention relates to a dry powdered LAMP rapid detection reagent and a rapid detection method based thereon, in particular to a dry powdered LAMP rapid detection kit for Streptococcus faecalis and a rapid detection method based thereon. Background technique [0002] Streptococcus faecalis is generally distributed in the intestinal tract of humans and animals and is excreted with feces. It is a thermostable bacterium that can still grow well at a temperature of 44.5±0.2°C. As drinking water, the water source is easily polluted directly or indirectly by feces. If there are intestinal infectious diseases in these pollution sources, the water source may contain live pathogenic factors. Contaminated drinking water will cause diseases in the crowd. As an indicator bacterium of sanitary bacteria in drinking water, it is very important in the detection of drinking water sanitation. Streptococcus faecalis can reflect whether the water body is polluted by fe...

AI Technical Summary

Problems solved by technology

In practical applications, the corresponding solution reaction system must be prepared before detection, and there are many components in it, which can easily lead to reagent contamination, detection interval contamination, and result errors caused by frequent operations. In addition, some reagents in the reaction system need to be frozen. , For example, the required Bst DNA polymerase needs to be stored strictly at -20°C. If the temperature is too high, it will affect its enzyme activity and cause the detection reagent to fail.

Therefore, the high-temperature long-distance transportation and on-site application of the reagent are inconvenient, and the corresponding cost is also increased.

At the same time, the current detection method has not been popularized in the fields of food safety and medicine, especially in the rapid detection of water bodies. As a non-standard method, the existing LAMP products are only for the detection process, and the methods of sample pretreatment are all no mention

Moreover, there are few kits for the rapid detection of Streptococcus faecalis based on loop-mediated isothermal amplification technology on the market, and similar dry powder products that can be used for normal temperature transportation have not yet appeared.

[0004] The main difficulties in the development of the dry powder type LAMP rapid detection kit are as follows: 1) Keep the activity of BstDNA polymerase during the freeze-drying process without loss; 2) Design suitable primers; 3) The color development is not obvious enough, making it difficult to distinguish the results

[0005] In order to maintain the activity of Bst DNA polymerase, adding a lyoprotectant is a more conventional choice. The commonly used lyoprotectant is trehalose, but its protective effect is relatively limited, and it is difficult to guarantee product quality

Images