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Method for rapidly extracting DNA of gram-positive bacterium genome based on paramagnetic particle method

A gram-positive bacteria and genome technology, applied in the biological field, can solve the problem of long extraction time, achieve the effects of economical extraction, simplify experimental steps and save extraction time

Inactive Publication Date: 2015-04-29
FUJIAN NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to provide a convenient, quick and practical method for quickly extracting the genomic DNA of gram-positive bacteria based on the magnetic bead method, and solve the problem that the extraction time required for the genomic DNA of gram-positive bacteria using conventional methods is long

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  • Method for rapidly extracting DNA of gram-positive bacterium genome based on paramagnetic particle method

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Embodiment 1

[0043] A method for rapidly extracting the genomic DNA of gram-positive bacteria based on a magnetic bead method, specifically comprising the following steps:

[0044] 1) Breakage of Gram-positive bacterial cells: Take 1ml of Staphylococcus overnight culture solution into a 1.5ml centrifuge tube, centrifuge at 12000rpm / min for 1min, discard the supernatant, add 1ml Staphylococcus overnight culture solution to the centrifuge tube, 12000rpm / min After centrifuging for 1-5 min, discard the supernatant, add 1ml of Staphylococcus overnight culture solution to the centrifuge tube, centrifuge at 12000rpm / min for 1min, discard the supernatant, (i.e. take 3ml of Staphylococcus overnight culture solution in total, and add in 3 times) into a 1.5ml centrifuge tube, centrifuge at 12000rpm / min for 1min each time and discard the supernatant), then add 0.25ml of double-distilled water to the bacterial pellet to suspend the pellet, and add 75μl of working solution A, incubate at 40°C for 30min, ...

Embodiment 2

[0058] A method for rapidly extracting the genomic DNA of gram-positive bacteria based on a magnetic bead method, specifically comprising the following steps:

[0059] 1) Breakage of Gram-positive bacterial cells: Take 1ml of Streptococcus overnight culture solution into a 1.5ml centrifuge tube, centrifuge at 10000rpm / min for 5min, discard the supernatant, add 1ml of Streptococcus overnight culture solution to the centrifuge tube, 10000rpm / min After centrifuging for 5 min, discard the supernatant, add 1ml of streptococcus overnight culture solution to the centrifuge tube, centrifuge at 10000rpm / min for 5min, discard the supernatant, (i.e., take 3ml of streptococcus overnight culture solution, and add it in 3 times. into a 1.5ml centrifuge tube, centrifuge at 10000rpm / min for 5min each time and discard the supernatant), then add 0.2ml of double distilled water to the bacterial pellet to suspend the pellet, and add 50μl of working solution A, incubate at 40°C for 10min, then add ...

Embodiment 3

[0072] A method for rapidly extracting genomic DNA of gram-positive bacteria based on a magnetic bead method, specifically comprising the following steps:

[0073] 1) Breakage of Gram-positive bacterial cells: Take 1ml of actinomycetes overnight culture solution into a 1.5ml centrifuge tube, centrifuge at 13000rpm / min for 2min, then discard the supernatant, add 1ml of actinomycetes overnight culture solution to the centrifuge tube, After centrifuging at 13000rpm / min for 2min, discard the supernatant, add 1ml of actinomycetes overnight culture solution to the centrifuge tube, discard the supernatant after centrifuging at 13000rpm / min for 2min, (that is, take a total of 3ml of actinomycetes overnight culture solution, divide into 3 Add it to a 1.5ml centrifuge tube once, centrifuge at 13000rpm / min for 2min each time and discard the supernatant), then add 0.5ml double distilled water to the bacterial pellet to suspend the pellet, add 80μl working solution A, and keep warm at 40°C ...

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Abstract

The invention discloses a method for rapidly extracting DNA of gram-positive bacterium genome based on a paramagnetic particle method. The method comprises the following three steps: breaking gram-positive bacterium cells, freeing DNA, extracting impurity protein, and rapidly purifying DNA of the genome by using the paramagnetic particle method. Compared with a conventional method for rapidly extracting the DNA of the gram-positive bacterium genome, the method is capable of not only saving a great deal of extraction time, but also simplifying the experiment steps, and the DNA of the gram-positive bacterium genome can be relatively effectively, rapidly and economically extracted.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a method for rapidly extracting genomic DNA of Gram-positive bacteria based on a magnetic bead method. Background technique [0002] Gram-positive bacteria are bacteria that can be stained dark blue or purple with Gram stain, while gram-negative bacteria cannot be stained (but are colored red for contrast). Gram-positive bacteria contain relatively large amounts of peptidoglycan in their cell walls, but often lack the second membrane and lipopolysaccharide layer possessed by Gram-negative bacteria. The cell wall of Gram-positive bacteria is relatively thick (20-80nm), mainly composed of peptidoglycan and teichoic acid, with only one layer of structure, and the main component is peptidoglycan, which accounts for 50%-90% of the dry weight of the cell wall . During the staining process, when treated with ethanol, the pore size in the network structure becomes small...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12R1/44C12R1/46
Inventor 王清水余彦
Owner FUJIAN NORMAL UNIV
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