Human feces DNA extraction method

An extraction method, feces technology, which is applied in the field of DNA extraction from human feces, can solve the problems of unforeseen and affecting the results of DNA extraction, and achieve the effect of high repeat stability

Active Publication Date: 2013-01-09
ZHEJIANG CENTURY CONDOR MEDICAL SCI & TECHNOLOTY CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are still problems in practical application: due to the particularity of fecal materials, relevant factors such as sample collection and storage will affect the results of DNA extraction (Frantzen et al. , 1998; Piggott and Taylor, 2003)
At present, domestic and foreign ...

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] 1. Stool sample collection:

[0036] Collect fecal samples excreted by people who have been exposed for no more than 24 hours; use selective samples to collect PSCs, scrape off the superficial mucosa of the fecal samples with a sterile blade, and divide them into 2ml centrifuge tubes;

[0037] 2. Sample storage:

[0038] Adopt low-temperature preservation method, collect the fecal samples, and immediately store them at -20°C;

[0039] 3. Sampling:

[0040] For the preserved feces samples, take them out and place them on ice, pick a 180mg sample with a sterilized pipette tip or a sterilized blade, and place them on ice;

[0041] 4. DNA extraction:

[0042] Extraction steps:

[0043] (1) Add 1.2ml of stool lysate into a 2ml sterilized Eppendorf tube, and place it on ice for operation;

[0044] (2) Weigh 180 mg of stool sample into each Eppendorf tube until the sample is fully lysed and mixed;

[0045] (3) Centrifuge at the highest speed for 5 minutes at room temperatu...

Embodiment 2

[0065] 1. Stool sample collection:

[0066] Collect fecal samples excreted by people who have been exposed for no more than 24 hours; use selective samples to collect PSCs, scrape off the superficial mucosa of the fecal samples with a sterile blade, and divide them into 2ml centrifuge tubes;

[0067] 2. Sample storage:

[0068] Using low-temperature preservation method, the collected fecal samples were immediately stored at -80°C;

[0069] 3. Sampling:

[0070] For the preserved feces samples, take them out and place them on ice, pick a 200mg sample with a sterilized pipette tip or a sterilized blade, and place them on ice;

[0071] 4. DNA extraction:

[0072] Extraction steps:

[0073] (1) Add 1.6ml of stool lysate into a 2ml sterilized Eppendorf tube, and place it on ice for operation;

[0074] (2) Weigh 200 mg of stool sample into each Eppendorf tube until the sample is fully lysed and mixed;

[0075] (3) Centrifuge at the highest speed for 5 minutes at room temperatu...

Embodiment 3

[0095] 1. Stool sample collection:

[0096] Collect fecal samples excreted by people who have been exposed for no more than 24 hours; use selective samples to collect PSCs, scrape off the superficial mucosa of the fecal samples with a sterile blade, and divide them into 2ml centrifuge tubes;

[0097] 2. Sample storage:

[0098] Adopt low-temperature preservation method, collect the feces samples and store them immediately at -50°C;

[0099] 3. Sampling:

[0100] For the preserved feces samples, take them out and place them on ice, pick a 190mg sample with a sterilized pipette tip or a sterilized blade, and place them on ice;

[0101] 4. DNA extraction:

[0102] Extraction steps:

[0103] (1) Add 1.4ml of stool lysate into a 2ml sterilized Eppendorf tube, and place it on ice for operation;

[0104] (2) Weigh 190 mg of stool sample into each Eppendorf tube until the sample is fully lysed and mixed;

[0105] (3) Centrifuge at the highest speed for 5 minutes at room temperat...

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PUM

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Abstract

The invention discloses a human feces DNA extraction method. The prior art is complicated and time-consuming; and the trace DNAs extracted by the previous sampling method is too little to perform molecular diagnosis. The extraction method uses selective sample collection PSC equipment for feces samples and adopts a low temperature preservation method. The extraction method comprises the following steps: picking a sample by using a sterilized gun head or sterilized blade and directly placing the picked sample on ice; and adding a feces lysis solution, fully cracking and evenly mixing, adding bovine serum albumin, centrifuging to remove the supernatant, adding protease K to perform a digestion reaction at 58 DEG C, then adding a protein precipitation solution to precipitate the protein, centrifuging to obtain the supernatant, adding a silica gel membrane binding solution to fully bind DNAs, centrifuging to obtain precipitates, adding a silica gel membrane washing solution to wash twice, centrifuging, and adding a TE (Tris + EDTA) buffer solution to finally obtain a preservation solution with DNAs. The human feces DNA extraction method has the advantages of high repeatability and stability, simpleness and short time.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a method for extracting DNA from non-damaging sampling materials, in particular to a method for extracting human feces DNA. Background technique [0002] Colorectal cancer is the most common digestive tract malignancy, second only to gastric cancer and esophageal cancer. The incidence of colorectal cancer has gradually increased in the past two decades, and at the same time, its age of onset tends to be aging. In western developed countries, colorectal cancer is the second malignant tumor after lung cancer. Some studies in recent years have shown that mutations in some genes such as K-ras, P53, APC, and MMR can lead to the transformation of normal colonic mucosal epithelial cells into tumor cells (Vogelstein, B. and Kinzler, K.W., 1993). Studies on these gene mutations Progress has made it possible to diagnose colorectal cancer through stool DNA testing. The feces of a normal person...

Claims

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Application Information

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IPC IPC(8): C12N15/10
Inventor 秦炜毕婷婷阮丽娟陈标榜余向东
Owner ZHEJIANG CENTURY CONDOR MEDICAL SCI & TECHNOLOTY CORP
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