Method for quickly extracting and preserving silica gel membrane type DNA
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A technology of silica gel membrane and room temperature storage, applied in the biological field, can solve the problems of high cost, inconvenient transportation of bacterial liquid or body fluid at low temperature, unusable places without laboratory conditions, etc., and achieve the effect of simple elution process
Inactive Publication Date: 2010-02-17
林桂兰
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The disadvantage of using method 1 is that laboratory equipment such as centrifuges and ultra-clean workbenches are required at the collection site, and this method cannot be used to extract and preserve DNA for samples collected in the field or places without lab
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Embodiment 1
[0020] 1.1. Take 6 grams of Trisbase, 8 grams of SDS, 1.5 grams of EDTA, and 80 grams of guanidine isothiocyanate in a 500ml volumetric flask, add 300ml of ultra-pure water, adjust the pH to 5.0 with concentrated hydrochloric acid, add deionized water to make it 500ml, Prepared as lysis buffer.
[0021] 2. Cut 1cm 2 Put the silica gel membrane into a plate, suck 200ul of the above-mentioned lysis buffer solution and drip it on the silica gel membrane, and let it dry in the air.
[0022] 3. Take 30ul of the bacteria solution that was enriched overnight and add it dropwise on the dried silica gel membrane, and let it dry at room temperature. Store the dried silica gel membrane in a plastic bag free from exogenous DNA contamination and keep it away from light.
Embodiment 2
[0024] 1. Take out the preserved silica gel membrane, put it into a 1.5ml centrifuge tube, add 1ml of 85% ethanol prepared with ultrapure water, place it on a horizontal shaker for 3 minutes, discard the ethanol, and discard the residual ethanol after a short centrifugation.
[0025] 2. Repeat step (1) to remove the salt and residual bacterial impurities on the silica gel membrane by washing twice.
[0026] 3. After short centrifugation, open the cap of the centrifuge tube and place it at room temperature for 10 minutes to remove ethanol.
[0027] 4. Add 100ul ultrapure water, place on a horizontal shaker for 10 minutes, and centrifuge briefly. The resulting aqueous solution contains bacterial genomic DNA and plasmid DNA, which can be used for subsequent sequencing, PCR, enzyme digestion, transformation and other reactions.
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Abstract
The invention discloses a method for quickly extracting and preserving a silica gel membrane type DNA. By utilizing the characteristic of a silica gel membrane that the silica gel membrane adsorbs DNAunder the conditions of high salt and low pH and releases the DNA under the conditions of low salt and high pH, the invention does not need a process of extracting the DNA in advance after the silicagel membrane is processed by a novel solution containing a cell cracking agent and a nuclease suppressor, cells are directly lysed, the DNA is adsorbed on the silica gel membrane, the silica gel membrane adsorbing the DNA can be preserved for a long time because of the existence of the nuclease suppressor and the degradation of the DNA is avoided. In the extracting process, the pure DNA is adsorbed on the membrane by the elution function of eluent I after salt and cell residues on the membrane are rinsed. By the elution function of eluent II, the DNA combined on the membrane is eluted into the solution. The whole DNA extracting and preserving process has quick operation and low price and is simple and convenient.
Description
technical field [0001] The invention relates to the field of biotechnology, in particular to a method for extracting and preserving DNA from bacteria or body fluids by using a silica gel membrane. Background technique [0002] At present, the technology of extracting DNA from bacteria or body fluids has been widely used, and there are various methods for extracting and preserving DNA. [0003] The traditional DNA extraction method is the phenol-chloroform method, which is cheap and widely used, but it is easy to cause environmental pollution due to the large amount of organic solvent used. The DNA extraction kit developed by using the DNA adsorption properties of resin, silica gel and silica gel membrane has little environmental pollution, but the operation steps are cumbersome, and the purified DNA needs to be obtained by multi-step centrifugation after the lysate is lysed. DNA cannot be preserved for long periods of time. [0004] The commonly used technology for DNA pre...
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