A primer composition for identifying intestinal microecological state and its application
A technology of intestinal microecology and primer set, which is applied in the field of primer composition composition and the prepared kit, and the primer composition for identifying the state of intestinal microecology, which can solve the problem of unpredictable overall situation of intestinal microecology, Strong limitations and other issues, to achieve the effect of reducing errors, reducing time costs, and high primer sensitivity
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Embodiment 1
[0126] The assembly of embodiment 1 kit
[0127] The design of primer composition, concrete sequence is as shown in table 1:
[0128] Table 1
[0129]
[0130]
[0131]
[0132] PCR reaction solution: Tris-HCl: 10-80mmol / L, pH8.5; Ammonium sulfate: 10-40mmol / L; MgCl 2 : 1-5.5mmol / L; dNTPs: 100-500μmol / L; final primer concentration: 0.25-1.25ng / μL. The components of the reaction solution in each reaction well are as follows:
[0133] Clostridium symbiotic PCR reaction solution: Tris-HCl: 10mmol / L, pH8.5; Ammonium sulfate: 10mmol / L; MgCl 2 : 3mmol / L; dNTPs: 200μmol / L;
[0134] Bifidobacterium adolescentis PCR reaction solution: Tris-HCl: 40mmol / L, pH8.5; Ammonium sulfate: 30mmol / L; MgCl 2 : 2.5mmol / L; dNTPs: 150μmol / L;
[0135] Fusobacterium nucleatum PCR reaction solution: Tris-HCl: 60mmol / L, pH8.5; Ammonium sulfate: 40mmol / L; MgCl 2 : 5.5mmol / L; dNTPs: 400μmol / L;
[0136] Anaerobic Peptostreptococcus PCR reaction solution: Tris-HCl: 80mmol / L, pH8.5; Ammonium s...
Embodiment 2
[0148] Embodiment 2 clinical verification
[0149] 1. Sample selection
[0150] 46 samples of intestinal microecological imbalance were collected in the hospital. All samples were confirmed by stool microscopy and doctors, including 8 cases of intestinal polyps and 38 cases of adenoma. At the same time, 28 healthy people with healthy physical examination were collected. The physical examination indicators were normal, and the fecal microscopy examination was normal. All of them had undergone colonoscopy examination and had no intestinal diseases.
[0151] 2. Genome extraction from stool samples
[0152] The bacterial genomes in different samples were extracted according to the instructions of the Fecal Bacterial Genome Extraction Kit (DP328, Tiangen Biochemical Technology (Beijing) Co., Ltd.), and the concentration and quality of DNA were detected.
[0153] 3. Check the samples to be tested
[0154] The extracted DNA was used as a template, and the loading amount was 5ng / sa...
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Abstract
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