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Feces preservation solution, preparation method thereof and feces preservation method

A technology for preserving liquid and feces, which is applied in biochemical equipment and methods, and the determination/inspection of microorganisms. safe effect

Pending Publication Date: 2019-07-12
康美华大基因技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The existing protective solution contains guanidine isosulfate cyanate, which has certain toxicity; or has a single component, which has a weak protective effect on cells

Method used

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  • Feces preservation solution, preparation method thereof and feces preservation method
  • Feces preservation solution, preparation method thereof and feces preservation method
  • Feces preservation solution, preparation method thereof and feces preservation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1. Sample source and preservation method

[0021] Fecal samples were collected from 3 volunteers. The samples of each volunteer were divided into 4 parts, and 4 different treatments were performed respectively, with 3 samples for each treatment:

[0022] (1) Freezing for one week: Weigh 0.2g of fecal samples, seal at -80°C, protect from light, store for one week, and then perform DNA extraction for future use;

[0023] (2) Preservation of formula 1 for one week: Weigh 0.2 g of fecal samples, use the control preservation solution formula (Formulation 1), seal at room temperature (25°C), protect from light, store for one week, and then perform DNA extraction for later use;

[0024] (3) Preservation of formula 2 for one week: Weigh 0.2g of fecal samples, use the preservation solution formula of the present invention (Formula 2), seal at room temperature (25° C.), protect from light, store for one week, and then perform DNA extraction for subsequent use;

[0025] ...

Embodiment 2

[0032] Example 2. Fecal sample processing method

[0033] The processing of the 4 stool samples from each of the above volunteers was as follows:

[0034] (1) Add 200 mg of zirconia to the stool sample tube and place on ice;

[0035] (2) Add 540μL SLX-Mlus Buffer (DNA extraction was done using Omega E.Z.N.A.Stool DNA Kit, Cat. No. D4015-02, the reagents for the extraction process are all from this kit), 60μL DS Buffer and 20μL Proteinase KSolution, put into a tissue grinder, 60HZ , 90s.

[0036] (3) 70°C, 500rpm metal bath incubation for 10min.

[0037] (4) Add 400 μL SP2Buffer, put it into a tissue grinder, 60HZ, 30s, and place on ice for 5min.

[0038] (5) Centrifuge at 20000g for 15min.

[0039] (6) Carefully pipette 800 μL of supernatant into a new 1.5 mL centrifuge tube.

[0040] (7) Add 400 μL of cHTR Reagent and vortex for 10s.

[0041] (8) After standing at room temperature for 2 minutes, centrifuge at 20000g for 2 minutes.

[0042] (9) Transfer 400 μL of supern...

Embodiment 3

[0054] Example 3. Detection of sample DNA concentration

[0055] The concentration of the extracted DNA samples was determined using a spectrophotometer. As shown in Table 2, the DNA concentration of formula 2 stored for one week is closer to the concentration of frozen storage for one week, while the concentration of formula 1 stored for one week is worse than that of formula 2 stored for one week and stored for one week at room temperature. This may be due to the fact that the composition of the flora has not been fixed at the initial state of sampling and has multiplied.

[0056] Table 2 Extraction product microplate reader detection result

[0057]

[0058]

[0059] A, B, and C represent 3 volunteers, 1 means cryopreservation method, 2 means formula 1 preservation, 3 means formula 2 preservation, 4 means room temperature preservation.

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Abstract

The invention provides a feces preservation solution, a preparation method thereof and a feces preservation method. The feces preservation solution contains 0.10-0.20 M of Tris base, 0.25-0.75 M of ethylenediaminetetraacetic acid (EDTA), 10 mM-room temperature saturated sodium chloride (NaCl), 5-15% of dimethyl sulfoxide (DMSO), 2.5-10% of ethanol and 5-20% of glycerol, wherein the pH of the fecespreservation solution is 7.0-9.0. The feces preservation solution can stably preserve microbial diversity in feces samples at normal temperature and improve DNA concentration, can ensure the accuracyof sequencing analysis of intestinal flora, realizes normal-temperature preservation and transportation of the feces samples, has a good preservation effect compared with an existing preservation solution, does not contain toxic components such as guanidine thiocyanate, and is safer to operate.

Description

technical field [0001] The present invention relates to a fecal preservation solution, a preparation method thereof, and a method for preserving feces. Background technique [0002] The human body contains a huge number of bacteria, even more than the number of human cells (10 13 ) is 10 times higher. These bacteria are distributed in the human intestinal tract and different tissues, and the intestinal tract, especially the large intestine, is the main site for bacterial colonization. The presence of these microbes has such a critical impact on human pathophysiology that researchers even call it an "organ" of the human body, with which they form a symbiotic relationship. Bacteria can not only ingest the food ingested by the human body, but also synthesize substances such as amino acids, vitamins and short-chain fatty acids, which are ingested by the human body, thereby promoting health. At the same time, pathogens or opportunistic pathogens can cause damage and cause dise...

Claims

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Application Information

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IPC IPC(8): C12Q1/6806
CPCC12Q1/6806C12Q2527/125
Inventor 杨晓春许冬瑾王苗苗胡悦
Owner 康美华大基因技术有限公司
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