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Isolation method and device for nucleic acid

A nucleic acid separation and nucleic acid technology, applied in the field of biochemistry, can solve the problems of poor nucleic acid separation effect and low recovery rate, and achieve the effects of high mechanical strength, improved recovery rate and easy processing.

Pending Publication Date: 2021-02-02
SHANGHAI GENECHEM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In view of the shortcomings of the prior art described above, the object of the present invention is to provide a nucleic acid separation method and device for solving the problems of poor nucleic acid separation effect and low recovery rate

Method used

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  • Isolation method and device for nucleic acid
  • Isolation method and device for nucleic acid
  • Isolation method and device for nucleic acid

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] Embodiment 1 is to the separation recovery effect of DNA ladder

[0091] Use a 6mm thick 3% agarose gel film to separate DNA ladders composed of DNA fragments of different lengths. Choose to use 40V, 80V, 120V, 160V, 200V and 240V voltage conditions, all energized for 90min, then recover DNA from the recovery compartment of the nucleic acid separation device, and perform 1% agarose gel electrophoresis analysis.

[0092] The result is as Figure 4 As shown, with the increase of voltage, the length of recovered DNA is getting longer and longer.

Embodiment 2

[0093] Example 2 to the separation effect of size continuous distribution DNA

[0094] Total DNA extracted from feces was separated using a 6 mm thick 3% agarose gel film. The voltage is 80V, energized for 20min, 40min, 60min and 90min respectively, and then the DNA is recovered from the recovery bin of the nucleic acid separation device. DNA was extracted using an equal volume of phenol: chloroform" isoamyl alcohol (25:24:1), added 10% volume of 3M NaAc (pH 5.2), added 2.5 volumes of ethanol, incubated at -20°C for 1 hour, and then centrifuged at 12000rpm The DNA precipitate was obtained. After the DNA was dissolved, 1 ng was taken, and the length was analyzed by capillary electrophoresis with BioAnalyzer2100.

[0095] The result is as Figure 5 and Figure 6 As shown, with the extension of the energizing time, the length of the obtained DNA becomes larger and larger.

Embodiment 3

[0096] Embodiment 3 is to the separation effect of size continuous distribution DNA

[0097] Escherichia coli (DL21) genomic DNA and total RNA were prepared. Take 130 μL of E.coli genomic DNA, fragmented to ~150bp and ~500bp respectively, and press E.coli complete genomic DNA: ~500bp interrupted genomic DNA: ~150bp interrupted genomic DNA: total RNA=2:1:1: Mix at a mass ratio of 0.1 to obtain a DNA test mixture. Purify the DNA test mixture obtained above using a 4mm thick 6% agarose gel film, power on at 80V for 10min, 15min, 20min, 30min and 40min, and recover the DNA in the recovery compartment of the nucleic acid separation device. Use the Qiagen gelextraction kit to recover the DNA from the film with the above electrification time, and then take the corresponding volume and perform 0.75% agarose gel electrophoresis to analyze the length of the DNA contained in the recovery chamber and the gel.

[0098] The result is as Figure 7 As shown, it shows that under the conditi...

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Abstract

The invention provides an isolation method for nucleic acid. The method at least comprises the following steps: agarose gel is put in an electrophoresis electric field, wherein the mass fraction of agarose in the agarose gel is 3%-6%; the width of the agarose gel in an electrophoresis direction is 4 mm-6 mm; agarose gel electrophoresis is performed on nucleic acid samples to be isolated, so that part of the nucleic acid is cut off in agarose gel, and part of nucleic acid moves out of the agarose gel to realize isolation of the nucleic acid. According to the isolation method for the nucleic acid, by utilizing high mechanical strength of high-concentration agarose, relatively thin gel can be supported; and the solid-liquid state of the agarose can be controlled through temperature, so that the thin gel is easy to process. The method can significantly improve the recovery rate of DNA. Sized isolation of the nucleic acid under the condition that the samples enter the gel not at the same time can be realized; electrophoresis abnormity during gel DNA overload can be effectively avoided through continuous sample loading; and the method can be used for isolating nucleic acid samples more than 1 mg.

Description

technical field [0001] The invention relates to the field of biochemistry, in particular to a nucleic acid separation method and device. Background technique [0002] Electrophoresis of nucleic acids is the most commonly used nucleic acid purification and size separation technique. The earliest electrophoresis of biomacromolecules was carried out in free solution, and then people found various electrophoretic support media, such as filter paper, cellulose acetate film, starch, agarose, polyacrylamide, etc. Currently, the main support media for nucleic acid electrophoresis are agarose and polyacrylamide. In molecular biology techniques, 0.5%-3% agarose gel is generally used, which can resolve DNA fragments of 80bp-4kb. Higher concentrations of agarose gels are also impractical with current molecular biology techniques. High-concentration agarose gel is easy to produce bubbles and dissolves unevenly during heating and dissolving; it solidifies quickly in the gel container u...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12M1/00C12M1/42
CPCC12N15/101
Inventor 吴帅来周雅李亚雷王庆亮朱向莹曹跃琼
Owner SHANGHAI GENECHEM