Escherichia coli and Shigella detection primers, kits and detection methods based on specific sequences

Abstract

The invention discloses escherichia coli and shigella detection primers based on a specific sequence, a kit and a detection method, belonging to the technical field of molecular biology. The primers are designed and synthesized for a specific sequence (as shown in SEQ ID NO. 3) shared by escherichia coli and shigella or a homologous sequence, the homology of which reaches 96% or above. The primers are specifically as shown in SEQ ID NO.1 and SEQ ID NO.2. The detection method comprises the steps of by taking genomic DNA of a to-be-detected sample or a single colony as a template, performing PCR amplification by means of the primers; judging the amplification primers as suspected positive if the amplification primers have same stripes with positive control; recovering the amplification primers which are suspected positive; performing sequencing and analyzing; and judging the primers which are as same as the sequence as shown in the SEQ ID NO.3 or the homology of which reaches 96% or above, as escherichia coli and shigella positive. According to the detection method, the operation is high, sensitivity and specificity are high, the result is consistent to species results of escherichia coli and shigella, the detection cost is low and the method has good promotion and application prospect.

Description

technical field [0001] The invention relates to a specific sequence-based detection primer for Escherichia coli and Shigella, as well as a kit containing the primer and a detection method for Escherichia coli and Shigella, belonging to the technical field of molecular biology. Background technique [0002] Escherichia coli (Escherichia coli) is one of the most common clinical pathogens, and it is also an important research object in the field of food hygiene and epidemiology in recent years. Since this bacterium often appears in places contaminated by feces, it is used as an indicator bacterium for fecal contamination of water sources and food. Although most Escherichia coli are facultative anaerobic resident flora in the intestinal tract of humans and animals, they are not pathogenic under normal circumstances, but some special serotypes of Escherichia coli are pathogenic. These pathogenic bacteria generally carry specific virulence genes, which can encode corresponding to...

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Problems solved by technology

[0004] At present, the detection methods of Escherichia coli and Shigella mainly include conventional detection methods and molecular biological detection methods, and conventional detection methods include morphological identification and biochemical test identification, among which morphological identification supplemented by biochemical tests is of great significance in bacterial identification , but this method also has some shortcomings, such as low recurrence rate, low discrimination ability, similar phenotypic characteristics cannot be equated with similar or closely related genotypes, etc.

Therefore, for those bacterial species that are difficult to distinguish through phenotypic characteristics, the conventional detection method cannot accurately identify the species, and the method is cumbersome to operate and the experimental period is long (Deng Meikui, Sun Ying, Han Wenqing. Bacterial identification methods [J]. Biomedical Advances in Engineering, 2014(02):84-88)

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