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Multiplex fluorescent PCR 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: 2018-05-29
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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  • Multiplex fluorescent PCR detection kit for periodontitis pathogenic bacteria
  • Multiplex fluorescent PCR detection kit for periodontitis pathogenic bacteria
  • Multiplex fluorescent PCR detection kit for periodontitis pathogenic bacteria

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] Example 1. Detection method of kit

[0083] 1.1 Sample collection: The sterile paper tip method is used to collect samples from the deepest part of the patient’s periodontal pocket or from the gingival sulcus of periodontal healthy persons. The specific collection method is as follows: After gargle with regular water, rinse with normal saline to remove food residues and blow Dry, moisturized cotton ball, topical disinfection with 2.5% iodine tincture cotton ball, insert a piece of sterile paper tip into the periodontal pocket or gingival sulcus, stop inserting when there is resistance, stay for 30s, take it out, and place it in a sterile centrifuge In the tube.

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

[0085] 1.3 Preparation of the reaction body fluid: the reaction volume is 20μL; each sample takes 3 clean 0.5ml EP tu...

Embodiment 2

[0093] Example 2. Sensitivity analysis of the kit

[0094] 2.1 Template: Synthesize plasmid DNA containing amplicons as a template for detection, use a micro-ultraviolet spectrophotometer to determine the concentration of purified plasmid DNA, calculate the number of copies of the initial DNA template by molecular weight, and then make 10-fold gradient dilutions in sequence Up to the number of copies, a total of 7 gradients, the number of copies: 1 x 10 6 Up to 1 x 10 0 .

[0095] 2.2 PCR detection: Use the above detection method to prepare the reaction solution, and then place the reaction tube on the fluorescent PCR machine for amplification and detection. After the reaction is completed, observe the fluorescence curve chart and analyze the minimum number of template copies that can be detected by the above kit. The specific amplification curve is as follows Figure 4 ~ Figure 13 Shown.

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

Embodiment 3

[0098] Example 3: Detection of clinical samples of the kit

[0099] 3.1 Sample collection: use the above-mentioned sample collection method to collect infection with Actinobacillus, Porphyromonas gingivalis, Prevotella intermedius, Treponema denticola, Forsysteinella, Fusobacterium nucleatum, Campylobacter gracilis, rectum Samples of 10 major pathogenic bacteria of periodontitis of Campylobacter, Eubacterium entangled and Streptococcus constellation, and sample DNA was extracted.

[0100] 3.2 PCR detection: Use the above detection method to prepare the reaction solution, and then place the reaction tube on the fluorescent PCR machine for amplification and detection.

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

[0102] by Figure 14 ~ Figure 23 It can be seen that using the above-mentioned PCR detection kit for periodontitis pathogens can detect all 10 main pathogens of periodontitis.

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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 flesh 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 disease has multiple and universal characteristics. The number of hospital visits for periodontitis patients nationwide is about 20 million per year; epidemiological studies conducted in the United States estimate that more than 4 million Americans suffer from periodontitis. Periodontitis is the result of multiple risk factors and susceptibility factors, which are intricate and affect each other. Risk factors such as the environment, behavior, and microorganisms directly increase the possibility of disease. A large number of studies have fo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/689C12Q1/686C12Q1/04C12R1/01
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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