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High-quality flora DNA extracting method

An extraction method and high-quality technology, applied in the field of DNA extraction, can solve the problems of cumbersome, foreign bacteria contamination, and complicated operation methods, and achieve the effect of good versatility, high concentration, and strong specificity

Inactive Publication Date: 2018-11-30
SHENZHEN LEADMED MEDICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But there are still the following problems: ①The generality of various extraction methods is not good
②Existing operating methods are too complex and cumbersome, and are easily contaminated by exogenous bacteria, resulting in the quality of extracted DNA not being guaranteed

Method used

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  • High-quality flora DNA extracting method

Examples

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Effect test

example 1

[0064] 1. Collection and storage of stool samples

[0065] Stool samples were collected using disposable sterile supplies at the time of sampling, and immediately after collection were stored separately in absolute ethanol and DETs.

[0066] 2. Sample pretreatment

[0067] Use a sterilized glass rod to mash the feces in the centrifuge tube into a paste, shake and mix well, take 2ml of the turbid liquid into a 2ml sterile centrifuge tube, centrifuge at 10,000rpm for 3min, and pour off the supernatant; Continue to add the turbid solution, centrifuge at 10000rpm for 3min to retain the precipitate, repeat this step 2-4 times to obtain about 200mg of feces precipitate for DNA extraction. Add 1ml of 0.9% physiological saline to the feces sediment, mix well and centrifuge at 10,000rpm for 3min, pour off the supernatant to retain the sediment, repeat this step twice, the purpose is to remove a small amount of preservation solution (absolute ethanol or DETs). solution, stored at roo...

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Abstract

The invention discloses a high-quality flora DNA extracting technique, which comprises four steps, namely sample collecting, preserving, pre-treatment and extracting, wherein in the step of extracting, firstly, a DNA self-sample is released from a lysis buffer solution containing cetyl trimethyl ammonium bromide; then, implementing separating and precipitating, so that a DNA precipitate is obtained; and finally, implementing adsorbing and purifying by virtue of guanidine thiocyanate and silicon dioxide, so that DNA, which is relatively high in purity, is obtained. The purity of the finished DNA prepared by the method is greater than 1.7 (OD260 / OD280), which is higher than that of a professional kit, and fragment is greater than 10kb, so that gene amplification and sequencing can be implemented. The method is simple and convenient to operate and easy for promotion and application; meanwhile, the method has the advantage of being good in universality; the high-quality DNA product can beobtained in various samples; and the method is applicable to related molecular experiment operations.

Description

technical field [0001] The invention relates to a DNA extraction method, in particular to a method for extracting high-quality bacterial flora DNA from human feces samples. Background technique [0002] Human activities are inseparable from microorganisms, and a large number of bacteria live in the human body. The intestinal micro-ecosystem is the largest micro-ecosystem in the human body, and it is estimated that there are about 10 14 An order of magnitude of normal flora is ten times the number of human cells. Intestinal flora is not only large in number and variety, but also closely related to human health. The research on flora has become an important topic in the field of life and health. Since the rise of fecal DNA analysis technology, great achievements have been made at home and abroad, and a variety of fecal DNA extraction methods have been developed, fully confirming the feasibility of fecal DNA analysis. But there are still the following problems: ① The general...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1003
Inventor 岳朋
Owner SHENZHEN LEADMED MEDICAL TECH
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