Multi-fluorescent PCR (polymerase chain reaction) detection kit for periodontitis pathogenic bacteria

A detection kit and multiple fluorescence technology, which are applied in the determination/inspection of microorganisms, microorganisms, biochemical equipment and methods, etc., can solve the problems such as the detection of periodontitis pathogens that have not yet occurred.

Active Publication Date: 2017-01-11
基音生物科技(衢州)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, there is no kit for the detection of periodontitis pathogenic bacteria on the market

Method used

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  • Multi-fluorescent PCR (polymerase chain reaction) detection kit for periodontitis pathogenic bacteria
  • Multi-fluorescent PCR (polymerase chain reaction) detection kit for periodontitis pathogenic bacteria
  • Multi-fluorescent PCR (polymerase chain reaction) detection kit for periodontitis pathogenic bacteria

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] Embodiment 1, kit detection method

[0083] 1.1 Collection of samples: Samples were collected from the deepest part of the periodontal pocket of the patient or the gingival sulcus of healthy periodontal patients using the sterile paper tip method. Dry, use cotton balls to separate moisture, sterilize the area with 2.5% iodine tincture cotton balls, insert a piece of sterile paper tip into the periodontal pocket or gingival sulcus, stop inserting when there is resistance, take it out after staying for 30 seconds, and place it in a sterile centrifuge tube.

[0084] 1.2 Preparation of sample DNA: The kit of the present invention does not provide a DNA sample extraction reagent, and the user can choose a suitable commercial kit to extract sample nucleic acid.

[0085] 1.3 Prepare the reaction body fluid: the reaction volume is 20 μL; take three clean 0.5ml EP tubes for each sample, respectively labeled as N-1, N-2, and N-3 (N is the sample number), and draw PCR 13 μL of b...

Embodiment 2

[0093] Embodiment 2, kit sensitivity analysis

[0094] 2.1 Template: Synthesize plasmid DNA containing amplicons as a template for detection, use a micro-ultraviolet spectrophotometer to measure the concentration of purified plasmid DNA, calculate the copy number of the initial DNA template by molecular weight, and then make 10-fold serial dilutions To the single digit copy number, a total of 7 gradients, the copy number is: 1 x 10 6 to 1 x 10 0 .

[0095] 2.2 PCR detection: The above detection method was used to prepare the reaction solution, and then the reaction tube was placed on a fluorescent PCR instrument for amplification and detection. After the reaction is completed, observe the fluorescence curve and analyze the minimum copy number of the template that can be detected by the above kit. The specific amplification curve is as follows: Figure 4 to Figure 13 shown.

[0096] 2.3 Results and analysis: by Figure 4 to Figure 13 It can be seen that the detection limit...

Embodiment 3

[0098] Embodiment 3, kit clinical sample detection

[0099] 3.1 Sample collection: Use the above-mentioned sample collection method to collect Actinomyces associated with infection, Porphyromonas gingivalis, Prevotella intermedia, Treponema denticola, Forsythia forsythia, Fusobacterium nucleatum, Campylobacter lenticularis, rectal Samples of 10 main pathogenic bacteria of periodontitis including Campylobacter, Eubacterium entanglement and Streptococcus constellation were collected, and DNA was extracted from the samples.

[0100] 3.2 PCR detection: the above-mentioned detection method is used to prepare the reaction solution, and then the reaction tube is placed on a fluorescent PCR instrument for amplification and detection.

[0101] 3.3 Results and analysis: After testing samples 1 to 3, the test results are as follows: Figure 14 ~ Figure 23 shown.

[0102] Depend on Figure 14 ~ Figure 23 It can be seen that all 10 main pathogenic bacteria of periodontitis can be detect...

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Abstract

The invention discloses a multi-fluorescent PCR (polymerase chain reaction) detection kit for periodontitis pathogenic bacteria. The multi-fluorescent PCR detection kit has the advantages that 10 types of main periodontitis pathogenic bacteria including actinobacillus actinomycetemcomitan, porphyromonas gingivalis, prevotella intermedia, treponema denticola, tannerella forsythia, fusobacterium nucleatum, campylobacter gracilis, campylobacter rectus, entanglement eubacterium and streptococcus constellatus can be detected by the multi-fluorescent PCR detection kit, the quality can be controlled in integral test procedures for acquiring samples, extracting nucleic acid, carrying out PCR amplification and the like, and detection results can be quickly obtained; the multi-fluorescent PCR detection kit for the periodontitis pathogenic bacteria can be used for quickly detecting the main periodontitis pathogenic bacteria.

Description

technical field [0001] The invention relates to a multiple fluorescent PCR detection kit for periodontitis pathogenic bacteria. Background technique [0002] Periodontitis is a multifactorial disease characterized by red and swollen gums, bleeding, pus, gingival meat atrophy, gingival leakage, and loose teeth. It is one of the common diseases in the human oral cavity. The national oral health epidemiological survey found that periodontitis is characterized by multiple occurrence and universality, and the number of patients with periodontitis in the country is about 20 million per year; epidemiological studies conducted in the United States estimate that more than 4 million patients with periodontitis Americans suffer from periodontal disease. Periodontitis is the result of multiple risk factors and susceptibility factors, which are intricate and interact with each other. Risk factors, such as environmental, behavioral, and microbial factors, directly increase the likelihoo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/04C12R1/01
CPCC12Q1/686C12Q1/689C12Q2600/16C12Q2537/143C12Q2563/107
Inventor 顾大勇沈澄
Owner 基音生物科技(衢州)有限公司
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