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Method for extracting nucleic acid from biological sample

A technique for extracting nucleic acids from biological samples, which is applied in the field of environmental microbiology to achieve the effect of wide coverage and easy operation

Inactive Publication Date: 2014-04-16
BGI SHENZHEN CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The development and use of various samplers has greatly promoted the development of air microbiology, but still no sampler is perfect

Method used

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  • Method for extracting nucleic acid from biological sample
  • Method for extracting nucleic acid from biological sample
  • Method for extracting nucleic acid from biological sample

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] 1. Preparation before sample collection

[0039] according to figure 1 C and figure 2 As shown in the structure, take a new cellulose acetate membrane, connect the filter collection assembly to the housing, and prepare the collection equipment.

[0040] 2. Sample collection

[0041] Turn on the switch of the air enrichment device, and under the pressure of 1000Pa<H≤3000Pa, start up and pump air continuously for 3 days in the workshops of the two breweries respectively. Inside, the filter membrane was torn off to collect the bacteria, and air sample 1 and air sample 2 were collected.

[0042] 3. Air sample nucleic acid extraction

[0043] Put the collected bacteria together with the torn surface filter membrane into a 50mL centrifuge tube, add 20mL TE, 200μL 10mg / mL lysozyme, 200μL 10mg / mL helicase and 200μL 10mg / mL lysozyme, blow Aspirate and mix well, incubate overnight at 37°C, digest with a mixer and mix evenly (overnight digestion); centrifuge the resulting di...

Embodiment 2

[0053] Put 0.5g of the collected bovine rumen sample into a 2mL centrifuge tube, add 500mL TE, 20μL 10mg / mL lysozyme, 20μL 10mg / mL helicase, 20μL 10mg / mL lysozyme, mix well by pipetting, and incubate overnight at 37°C , digest with a mixer (overnight digestion); centrifuge the resulting digestion solution at 4°C at 9000r / min for 10 minutes, remove the supernatant, and keep the precipitate; add 400 μ TENS lysate (200 mmol / L NaCl, 100mmol / L Tris-HCl, pH 8.0, 2.0%SDS, 50mmol / L EDTA, 0.5%Triton X-100) and 20mg / mL proteinase K 300μL, invert and mix well, incubate at 55℃ for 1h, invert and mix every 15min Homogenize once; Centrifuge the lysed sample at 10000r / min at 4°C for 10min, keep the supernatant, discard the precipitate; add an equal volume of phenol:chloroform:isoamyl alcohol (25:24:1) to the supernatant , mix well, and centrifuge at 12000r / min at 4°C for 10min; transfer the obtained supernatant to a new tube, add an equal volume of chloroform:isoamyl alcohol (24:1) to it, mi...

Embodiment 3

[0055] Adopt dental plaque sample, repeat embodiment 2 to extract DNA, gained experimental result sees Figure 8 .

[0056] like Figure 7 and Figure 8 As shown, using the method of the present invention, nucleic acid samples can be effectively extracted from microorganisms.

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Abstract

The invention provides a method for extracting nucleic acid from a biological sample. The method for extracting nucleic acid from biological samples comprises the following steps: cracking the biological sample to obtain a cracking product containing nucleic acid; and separating nucleic acid from the cracking product. The cracking of the biological sample further comprises: digesting the biological sample by a mixture containing lysozyme, helicase, and lyticase; centrifuging the digested product, collecting the precipitate; cracking the precipitate by a lysate to obtain the cracking product containing nucleic acid. The method can effectively extract nucleic acid from a biological sample.

Description

technical field [0001] The invention relates to environmental microbiology, in particular, the invention relates to methods for collecting microorganisms in fluids and extracting nucleic acids. Background technique [0002] Air is the environment in which human beings live, and it is also the medium for microorganisms to spread and spread diseases. Microbes in the air mainly include bacteria, fungi, viruses and actinomycetes, which are the main components of air pollution. closely related to human activities. Microorganisms in the air mainly come from water, soil, animals, plants and humans in nature, agricultural activities, animal breeding, industrial production, sewage treatment, fermentation process and food production plants, etc. The number of microorganisms in the air is one of the important standards of air quality . To understand the microorganisms in the air, it is necessary to enrich the microorganisms for observation and analysis, which requires a specially des...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
Inventor 郭晶章文蔚肖亮李子华
Owner BGI SHENZHEN CO LTD
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