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Magnetic bead nucleic acid extraction method

An extraction method and a technology of magnetic bead method, which are applied in the fields of biochemical equipment and methods, DNA preparation, and microbial determination/inspection, etc., and can solve the problems of incomplete separation, easy loss of nucleic acid DNA/RNA, low DNA purity and concentration, etc. , to achieve the effect of improving concentration and purity

Inactive Publication Date: 2017-04-26
解码(上海)生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In the prior art, the magnetic bead nucleic acid extraction method requires multiple elutions and washings during the extraction process, and the nucleic acid DNA / RNA is easily lost during the extraction process, which affects the purity and concentration of the final extracted nucleic acid DNA / RNA
After adding washing buffer to the magnetic bead-nucleic acid complex, when washing to remove impurities on the magnetic bead-nucleic acid complex, it is necessary to apply the effect of an external magnetic field, which may easily cause incomplete separation of the washing liquid and the magnetic bead-nucleic acid complex. , the DNA purity and concentration are low, and the final extracted sample cannot fully meet the requirements of the test

Method used

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Embodiment 1

[0024] A method for extracting nucleic acid by magnetic bead method, comprising the following steps: S1, putting 200 μl of saliva biological sample into a centrifuge tube, adding 1000 μl of lysis buffer solution with a pH value of 7.4 to the centrifuge tube, mixing at room temperature for 3 minutes, and then centrifuging The tube is placed on a magnetic stand, magnetically separated for 20s, and the nucleic acid DNA / RNA in the biological sample is lysed and separated, and the nucleic acid DNA / RNA is combined with the nano-magnetic beads with a diameter of 20nM in the lysis buffer, and the nano-magnetic The beads are superparamagnetic silicon oxide nano-magnetic microbeads, which form magnetic bead-nucleic acid complexes under the action of an external magnetic field; discard the supernatant;

[0025] Wherein, of lysis buffer; The composition of lysis buffer is: 2M guanidine hydrochloride, 2M sodium iodide and 10mM EDTA; 1% Triton X-100 (Triton X-100), 1% Tween-20 (Tween- 20), ...

Embodiment 2

[0031] A method for extracting nucleic acid by magnetic bead method, comprising the following steps: S1, putting 200 μl of saliva biological sample into a centrifuge tube, adding 1000 μl of lysis buffer solution with a pH value of 7.4 to the centrifuge tube, mixing at room temperature for 3 minutes, and then centrifuging The tube is placed on a magnetic stand, magnetically separated for 20s, and the nucleic acid DNA / RNA in the biological sample is lysed and separated, and the nucleic acid DNA / RNA is combined with the nano-magnetic beads with a diameter of 20nM in the lysis buffer, and the nano-magnetic The beads are superparamagnetic silicon oxide nano-magnetic microbeads, which form magnetic bead-nucleic acid complexes under the action of an external magnetic field; discard the supernatant;

[0032] Wherein, of lysis buffer; The composition of lysis buffer is: 2M guanidine hydrochloride, 2M sodium iodide and 10mM EDTA; 1% Triton X-100 (Triton X-100), 1% Tween-20 (Tween- 20), ...

Embodiment 3

[0038] A method for extracting nucleic acid by magnetic bead method, comprising the following steps: S1, putting 200 μl of saliva biological sample into a centrifuge tube, adding 1000 μl of lysis buffer solution with a pH value of 7.4 to the centrifuge tube, mixing at room temperature for 3 minutes, and then centrifuging The tube is placed on a magnetic stand, separated by magnetic force for 20 s, and the supernatant is discarded; the nucleic acid DNA / RNA in the biological sample is lysed and separated, and the nucleic acid DNA / RNA is mixed with the nano-magnet with a diameter of 20 nM in the lysis buffer. Bead binding, nano-magnetic beads are superparamagnetic silica nano-magnetic micro-beads, which form magnetic bead-nucleic acid complexes under the action of an external magnetic field;

[0039] Wherein, of lysis buffer; The composition of lysis buffer is: 2M guanidine hydrochloride, 2M sodium iodide and 10mM EDTA; 1% Triton X-100 (Triton X-100), 1% Tween-20 (Tween- 20), 1% ...

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Abstract

The invention discloses a magnetic bead nucleic acid extraction method. The method includes: adding a cracking buffer solution into a biological sample, and conducting cracking separation of nucleic acid in the biological sample; combining nucleic acid with nano magnetic beads in the cracking buffer solution to form a magnetic bead-nucleic acid compound under the action of an external magnetic field; adding a washing buffer solution into the magnetic bead-nucleic acid compound to conduct centrifugal washing so as to remove impurities from the magnetic bead-nucleic acid compound, and collecting the washed magnetic bead-nucleic acid compound; adding an elution buffer solution into the washed magnetic bead-nucleic acid compound, and adding a disinhibition agent; directly subjecting a mixture of the magnetic bead-nucleic acid compound and the elution buffer solution to subsequent fluorescence quantitative PCR reaction. The method provided by the invention replaces a magnetic frame with centrifugation for adsorption of magnetic beads, and separates eluent and magnetic beads, can set different rotation speeds and time according to different types of nucleic acid so as to thoroughly separate the eluent and magnetic beads, thus improving the extraction concentration and purity.

Description

technical field [0001] The invention belongs to the technical field of nucleic acid extraction, in particular to a magnetic bead nucleic acid extraction method. Background technique [0002] The magnetic bead method for nucleic acid extraction is to use nanotechnology to improve and modify the surface of superparamagnetic nanoparticles, and then prepare superparamagnetic silica nano-magnetic beads. The magnetic beads can specifically recognize and efficiently combine with nucleic acid molecules on the microscopic interface. Utilizing the superparamagnetism of silicon oxide nanospheres, under the action of Chaotropic salt (guanidine hydrochloride, guanidine isothiocyanate, etc.) After separation, it can be used in various fields such as clinical disease diagnosis, blood transfusion safety, forensic identification, environmental microbial detection, food safety detection, and molecular biology research. [0003] Magnetic bead nucleic acid extraction can generally be divided ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/68
CPCC12N15/1013C12Q1/6806C12Q2563/143C12Q2563/149
Inventor 徐赛涛吴勇聪温小媛陶瑞
Owner 解码(上海)生物医药科技有限公司
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