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Primer probe group, kit and detection method for detecting fusobacterium nucleatum in saliva

A Fusobacterium nucleatum, primer probe technology, applied in the field of microbial molecular biology detection, can solve the problem of low accuracy of Fusobacterium nucleatum, achieve the effect of saving money, saving time, and improving detection efficiency

Pending Publication Date: 2020-04-10
SHANDONG UNIV QILU HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

This patented technology allows us to accurately measure bacillary acid production by measuring specific enzyme activity within different parts of our body called nucleuses or cells that produce it. By comparing this measurement data against previous measurements from similar individuals we aimed at finding out any deviations between these two values during disease diagnosis. These findings could help make medical decisions based on factors like drug use, diet regimen changes, etc., resulting in improved patient care quality and reduced costs associated therewith.

Problems solved by technology

The technical problem addressed in this patented text relates to developing a tool called fuzzy genetic analysis (FCA) that allows clinicians to quickly identify specific types of organisms from their blood serums without being influenced by extracting pure nucleic acid samples.

Method used

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  • Primer probe group, kit and detection method for detecting fusobacterium nucleatum in saliva
  • Primer probe group, kit and detection method for detecting fusobacterium nucleatum in saliva
  • Primer probe group, kit and detection method for detecting fusobacterium nucleatum in saliva

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Effect test

Embodiment 1

[0088]1. Extract the DNA in the saliva of 20 cases of healthy persons undergoing colonoscopy, patients with intestinal hyperplastic polyps, patients with adenoma, and patients with colorectal cancer. There was no significant difference in gender and age among the groups (P>0.05).

[0089] 2. 6 commonly used internal reference genes (GAPDH, RPPH1, NAGK, TERT, ERV-3, SLCO2A1) were detected by qPCR. After evaluation by NormFinder and geNorm software, it was found that the combination of GAPDH and TERT has a better effect on DNA quantification in saliva samples. Stability, see Table 1.

[0090] Table 1 Stability evaluation of internal reference genes

[0091]

[0092] 3. Design primers and fluorescent probes for Fusobacterium nucleatum DNA and internal reference genes GAPDH and TERT. According to the TaqMan fluorescent probe PCR detection method, determine the design principles of primers and fluorescent probes to avoid mutual interference and interference between each primer ...

Embodiment 2

[0100] Case data: Saliva samples were collected from 20 healthy persons undergoing colonoscopy, patients with intestinal hyperplastic polyps, patients with adenoma, and patients with colorectal cancer. There was no significant difference in gender and age among the groups (P>0.05).

[0101] (3) Collection of serum samples: Use a saliva collection tube to collect the non-irritating whole saliva secreted by the patient's mouth after gargling with water, centrifuge at 3000rpm for 10min, then centrifuge at 11000rpm for 10-15min, collect the supernatant, and store it at -80°C.

[0102] (4) Extract DNA from saliva: Take about 200 μL saliva, add 20 μL proteinase K, 37 ° C for 1-2 hours; then add 200 μL buffer AL, vortex mix for 15 seconds, incubate at 56 °C for 10 min, add 200 μL absolute ethanol, vortex mix After standing still for 15s, vortex again for 15s, centrifuge, transfer the resulting mixture to a QLAamp micro spin column (in a 2mL collection tube), centrifuge at 6000g for 1...

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Abstract

The invention provides a primer probe group, a kit and a detection method for detecting fusobacterium nucleatum in saliva, belonging to the technical field of microbial molecular biological detection.The primer probe group comprises the following components: a fusobacterium nucleatum DNA upstream primer, a fusobacterium nucleatum DNA downstream primer, a fusobacterium nucleatum DNA probe, a GAPDHupstream primer, a GAPDH downstream primer, a GAPDH probe, a TERT upstream primer, a TERT downstream primer and a TERT probe. The primer probe group can be used for detecting the DNAs and the reference genes of fusobacterium nucleatum in saliva at the same time, and correcting fusobacterium nucleatum DNA detection deviation through the correction of the reference genes, thereby achieving the purpose of accurate quantification. Besides, compared with other reference gene combinations, the combination of GAPDH and TERT has good stability in DNA quantification of saliva samples, and provides guarantee for accurate quantification of target molecules.

Description

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Claims

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Application Information

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Owner SHANDONG UNIV QILU HOSPITAL
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