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Method for separating and purifying pathogenic microorganism DNA

A pathogenic microorganism and purification column technology, applied in the field of DNA extraction, can solve the problems of insufficiency, low load of pathogenic microorganisms, long time consumption, etc., and achieve the effects of improving work efficiency, high content and purity, and simple and quick operation process.

Pending Publication Date: 2020-06-05
广州达正生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, when sequencing the pathogenic microorganisms infected in a low amount in human or animal samples, we are faced with such a problem: because the load of pathogenic microorganisms in infected samples is very small, it is necessary to perform ultra-high-depth sequencing on infected samples in order to Obtain information on low loads of infected pathogenic microorganisms
The reagent consumables used in this method are expensive and time-consuming, which cannot meet the needs of clinical specimen testing

Method used

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  • Method for separating and purifying pathogenic microorganism DNA
  • Method for separating and purifying pathogenic microorganism DNA
  • Method for separating and purifying pathogenic microorganism DNA

Examples

Experimental program
Comparison scheme
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Embodiment 1

[0044] 1. Reagent kit:

[0045]Nucleic acid release and binding agent: containing 3M GuSCN, 16.7% V / V ethanol, 2% V / V Triton X100, 10mM EDTA, 5mM Tri-HCl, mass fraction 0.5% SDS, and the solvent is water. Preparation method: weigh each component according to its content, then mix the components except ethanol and Triton X100 evenly, and then sterilize. The sterilization method is 121℃104.3KPa for 15 minutes, and then add ethanol and Triton X100.

[0046] Nucleic acid purification:

[0047] Wash solution 1: containing 3M GuSCN, 16.7% V / V ethanol, 2% V / V Triton X100, 10 mM EDTA, 5 mM Tri-HCl, and the solvent is water. Preparation method: weigh each component according to its content, then mix the components except ethanol and Triton X100 evenly, and then sterilize. The sterilization method is 121℃104.3KPa for 15 minutes, and then add ethanol and Triton X100.

[0048] Washing solution 2: containing 25% ethanol (V / V), 100 mM sodium chloride, 10 mM Tri-HCl, and the solvent is w...

Embodiment 2

[0094] This embodiment is the same as Embodiment 1, except:

[0095] The nucleic acid release and binding agent includes the following components: 2M GuSCN, 15% V / V ethanol, 1% V / V TritonX100, 8mM EDTA, 3mM Tri-HCl, 0.3% SDS by mass fraction, and the solvent is water.

[0096] The washing liquid 1 includes: 2M GuSCN, 15% V / V ethanol, 1% V / V Triton X100, 8mM EDTA, 3mM Tri-HCl, and the solvent is water;

[0097] The washing liquid 2 includes: 20% V / V ethanol, 80 mM sodium chloride, 8 mM Tri-HCl, and the solvent is water.

Embodiment 3

[0099] This embodiment is the same as Embodiment 1, except:

[0100] The nucleic acid release and binding agent includes the following components: 4M GuSCN, 17% V / V ethanol, 3% V / V TritonX100, 11 mM EDTA, 6 mM Tri-HCl, 0.6% SDS by mass fraction, and the solvent is water.

[0101] The washing solution 1 includes: 4M GuSCN, 17% V / V ethanol, 3% V / V Triton X100, 11mM EDTA, 6mM Tri-HCl, and the solvent is water;

[0102] The washing liquid 2 includes: 30% V / V ethanol, 120 mM sodium chloride, 11 mM Tri-HCl, and the solvent is water.

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Abstract

The invention discloses a method for separating and purifying pathogenic microorganism DNA. The method comprises the steps of transferring samples into a centrifugal tube, adding nucleic acid releasing and binding agents in claim 1 or 2, performing uniform vortex mixing, performing incubation, then performing transferring into a purification column, performing centrifuging to remove filtrate, loading the purification column back into a collecting tube, and performing washing and eluting to obtain the pathogenic microorganism DNA. The method special for separating and purifying the pathogenic microorganism DNA disclosed by the invention is simple, convenient and quick in operation process, enough DNA can be obtained through a few samples, and the separated DNA is high in content and purity.

Description

Technical field: [0001] The invention belongs to the field of DNA extraction, and in particular relates to a method for separating and purifying DNA of pathogenic microorganisms. Background technique: [0002] In the field of molecular detection in the past ten years, high-throughput sequencing technology has developed rapidly. It can perform parallel sequencing of millions to billions of DNA molecules at a time, which makes it possible to perform in-depth and detailed analysis of the transcriptome and genome of a species. , Overall analysis. The unique advantages of high-throughput sequencing technology not only promote the progress of scientific research, but also show great application value in the field of clinical medicine. [0003] However, when sequencing the pathogenic microorganisms infected in a low amount in human or animal samples, we are faced with such a problem: since the load of pathogenic microorganisms in infected samples is very small, it is necessary to ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6806
CPCC12Q1/6806C12Q2523/308C12Q2527/125
Inventor 杨俊范明姣唐续赵应洪
Owner 广州达正生物科技有限公司
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