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Magnetic bead-based DNA extraction method, lysate and kit

An extraction method and lysate technology, which are applied in the field of magnetic bead-based DNA extraction methods and lysates, can solve the problems of inability to achieve complete mechanized treatment, time-consuming and labor-intensive, etc., and achieve the reduction of liquid addition operation steps, convenient use and high safety. Effect

Active Publication Date: 2018-12-18
武汉纳磁生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In view of the above defects or improvement needs of the prior art, the present invention provides a DNA extraction method, lysate and kit based on magnetic beads. Optimize the experimental steps of DNA extraction, and design the corresponding kit to realize the whole process of DNA extraction in one liquid addition operation, and realize the whole process of mechanized processing, which is suitable for the existing magnetic bead method DNA extractor, thus solving the existing magnetic bead-based DNA extraction instrument. The current DNA extraction method cannot achieve complete mechanized processing, time-consuming and labor-intensive technical problems

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  • Magnetic bead-based DNA extraction method, lysate and kit
  • Magnetic bead-based DNA extraction method, lysate and kit

Examples

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Effect test

Embodiment 1

[0056] A kit for cooperating with the magnetic rod method nucleic acid automatic extraction instrument, including 200ul of lysate, 15ul of proteinase K solution with a concentration of 10mg / ml, 600ul of magnetic bead dispersion with a concentration of 4mg / ml, 400ul of the first washing solution, and 400ul of the second washing solution. Washing solution 500ul, third washing solution 500ul, and eluent 30ul;

[0057] The pH value of the lysate is 7 buffer, containing 140mmol NaCl, 3mol / L chaotropic agent, 5mmol EDTA, 5% polyethylene glycol 6000, and 2% Triton. The chaotropic agent is guanidine hydrochloride.

[0058] The particle size of the magnetic beads in the magnetic bead dispersion is 400 mm.

[0059] The pH value of the first washing solution is 7.8, containing 100mmol / L NaCl, 5mmol / l EDTA, 10mmol / l Tris, 0.5% Triton by mass fraction, and 0.4% SDS by mass fraction;

[0060] The second washing solution contains 100mmol / L NaCl and 70% alcohol by mass fraction;

[0061] T...

Embodiment 2

[0081] A kit for cooperating with the magnetic rod method nucleic acid automatic extraction instrument, including 300ul of lysate, 10ul of proteinase K solution with a concentration of 20mg / ml, 500ul of magnetic bead dispersion with a concentration of 6mg / mL, 500ul of the first washing solution, and 500ul of the second washing solution. Washing solution 600ul, third washing solution 600ul, and eluent 50ul;

[0082] The pH value of the lysate is 8, containing 140mmol NaCl, 5mol / L chaotropic agent, 10mmol EDTA, 10% polyethylene glycol 8000, and 2% Triton. The chaotropic agent is guanidine isothiocyanate.

[0083] The particle size of the magnetic beads in the magnetic bead dispersion is 400mm.

[0084] The pH value of the first washing solution is 8.0, containing 200mmol / L NaCl, 10mmol / l EDTA, 25mmol / l Tris, 1% Triton by mass fraction, and 0.5% SDS by mass fraction;

[0085] The second washing solution contains 200mmol / L NaCl and 75% alcohol by mass fraction;

[0086] The thi...

Embodiment 3

[0106] A kit for cooperating with the magnetic rod method nucleic acid automatic extraction instrument, including 250ul of lysate, 5ul of proteinase K solution with a concentration of 30mg / ml, 400ul of magnetic bead dispersion with a concentration of 8mg / ml, 600ul of the first washing solution, and 600ul of the second washing solution. Washing solution 700ul, third washing solution 700ul, and eluent 100ul;

[0107] The lysate is a buffer solution with a pH value of 9, containing 200 mmol of NaCl, 6 mol / L chaotropic agent, 20 mmol of EDTA, 15% polyethylene glycol 10000 by mass fraction, and 4% Triton by mass fraction. The chaotropic agent is guanidine hydrochloride.

[0108] The particle size of the magnetic beads in the magnetic bead dispersion is 400mm.

[0109] The pH value of the first washing solution is 8.5, containing 250mmol / L NaCl, 20mmol / l EDTA, 30mmol / l Tris, 4% Triton by mass fraction, and 2% SDS by mass fraction;

[0110] The second washing solution contains 250m...

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Abstract

The invention discloses a magnetic bead-based DNA extraction method, a lysate and a kit. The lysate is characterized in that a buffer solution with a sodium ion content of greater than or equal to 140mmol and a pH value of 7 to 9 contains a chaotropic agent with a concentration of 3 mol / L to 6 mol / L, 5 mmol to 20 mmol of a metal chelating agent and 5% to 15% by mass of polyethylene glycol. The extraction method comprises the following steps: (1) acquiring a cleavage reaction system; (2) treating the cleavage reaction system; (3) adding nanometer magnetic beads; (4) acquiring magnetic beads binding to DNAs; (5) carrying out washing and then performing DNA elution. The kit comprises the lysate, a proteinase K solution, a magnetic bead dispersion liquid, a first washing solution, a second washing solution, a third washing solution and an eluent. Reagents used in the invention are non-toxic, so experimental safety is improved; and only one-step liquid adding operation is required, so labor cost is greatly reduced.

Description

technical field [0001] The invention belongs to the field of nucleic acid extraction, and more specifically relates to a magnetic bead-based DNA extraction method and a lysate. Background technique [0002] The DNA extraction method is the basis of DNA monitoring and has important application prospects. At present, the biological samples for DNA extraction are hair roots, oral epithelial cells, etc.; oral epithelial cells are composed of multiple layers of cells, and the basal cells can continuously divide and proliferate. To replenish the aging or damaged cells on the surface, it has strong regeneration ability, strong metabolism, and easy to fall off. Epithelial cells have complete genomic DNA like other types of cells. Extracting oral epithelial cells is a simple, non-invasive and painless sampling method, suitable for people of all ages. Therefore, it is often used as an experimental material for disease diagnosis and scientific research. [0003] Traditional DNA extra...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013C12Q2521/537C12Q2527/125
Inventor 冯思文吴晗彬孙祥忠王佳港
Owner 武汉纳磁生物科技有限公司
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