Magnetic bead mixture, nucleic acid preservation method, nucleic acid extraction reagent and kit

A mixed liquid and magnetic bead technology, applied in the field of biochemistry, can solve the problems of inability to store at room temperature or refrigeration, inability to judge DNA fragmentation, complicated preparation, etc., to reduce packaging costs and transportation costs, save sample addition steps, Simple to use effects

Active Publication Date: 2021-08-17
FOSHAN UNITED MEDICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0008] At present, the Chinese patent application CN102559654A discloses a DNA preservation solvent, the solvent components include glycerol, TE buffer, 1-carboxy-N,N,N-trimethylbetaine, this DNA preservation solution has more components , the preparation is complicated and the cost is high
Another Chinese patent application CN106434636A discloses a novel DNA stabilization solution and its preparation method. The solution formula mainly includes potassium phosphate, glucose, disodium edetate and water, which can be stored for more than one year without degradation by agarose gel electrophoresis detection. , but based on agarose detection, it is impossible to judge the fragmentation of DNA and this method needs to be stored at -20°C or -80°C, and cannot be stored at room temperature or refrigerated

Method used

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  • Magnetic bead mixture, nucleic acid preservation method, nucleic acid extraction reagent and kit
  • Magnetic bead mixture, nucleic acid preservation method, nucleic acid extraction reagent and kit
  • Magnetic bead mixture, nucleic acid preservation method, nucleic acid extraction reagent and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] 1. The nucleic acid extraction steps are as follows:

[0056] (1) Add 450 μL of cell lysate, 40 μL of magnetic bead mixture, and 250 μL of sample into a 1.5 mL centrifuge tube, mix well for 2 minutes, and then let stand at room temperature for 10 minutes. Perform a brief centrifugation to absorb the magnetic beads and discard all the solution.

[0057] (2) Add 500 μL of washing solution, mix for 2 minutes, and then let stand at room temperature for 2 minutes. Perform a brief centrifugation to absorb the magnetic beads and discard all the solution.

[0058] (3) Add 500 μL of washing solution, mix for 2 minutes, and then let stand at room temperature for 2 minutes. Perform a brief centrifugation to absorb the magnetic beads and discard all the solution.

[0059] (4) Dry the magnetic beads at room temperature for 10 minutes, add 80 μL of eluent, mix for 2 minutes, and then let stand at room temperature for 5 minutes. Perform brief centrifugation to absorb the magnetic ...

Embodiment 2

[0083] The nucleic acid extraction step and fluorescent quantitative PCR detection step of Example 1 were used to analyze the recovery rate of the magnetic bead mixture.

[0084] The magnetic bead mixed liquid formula of the present embodiment is as described in table 6:

[0085] Table 6

[0086] component name Dosage range per mL magnetic beads 100mg Proteinase K 50mg urea 10mM Tris-Hcl 100mM CaCl 2

10mM EDTA-Na 2

10mM glycerin 50％ Carrier RNA 25μg DNase and RNase free purified water Replenish to 1mL

[0087] Detection of the recovery rate of the magnetic bead mixture:

[0088] a) Use negative serum to dilute the recombinant plasmid to 5.6×10 4 copy / μL, the magnetic bead mixture was used for extraction, and the PCR reaction reagent was used for detection.

[0089] b) Dilute the recombinant plasmid to 1.75×10 with DNase / RNase-free purified water 5 copy / μL, detected by PCR reaction reagent...

Embodiment 3

[0099] The nucleic acid extraction step and fluorescent quantitative PCR detection step of Example 1 were used to analyze the recovery rate of the magnetic bead mixture.

[0100]The magnetic bead mixed liquid formula of the present embodiment is as described in table 8:

[0101] Table 8

[0102]

[0103]

[0104] Detection of the recovery rate of the magnetic bead mixture:

[0105] a) Dilute the recombinant plasmid to 1×10 with negative serum 5 copy / μL, 1×10 4 copy / μL, 1×10 3 copy / μL, the magnetic bead mixture is used for extraction, and the reaction reagent is used for detection.

[0106] b) Use DNase / RNase-free purified water to dilute the recombinant plasmid to 3.125×10 5 copy / μL, 3.125×10 4 copy / μL, 3.125×10 3 copy / μL, directly detected by the reaction reagent.

[0107] Table 9

[0108]

[0109]

[0110] c) The results are shown in Table 9 and image 3 As shown, after the mixed recombinant plasmid serum was extracted, 1×10 5 copy / μL (equivalent to 3....

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Abstract

The invention discloses a magnetic bead mixed liquid, which comprises magnetic beads, protease, denaturing agent, glycerin, buffer and nucleic acid sedimentation aid. The invention also discloses a nucleic acid preservation method, a nucleic acid extraction reagent and a kit. The components of the magnetic bead mixture are premixed in a certain proportion. When nucleic acid extraction is performed, the corresponding volume can be taken at one time to add the required components evenly. The dosage is more accurate and the operation is simpler; the components are premixed in proportion. Can reduce packaging cost and transportation cost. The magnetic bead mixture solution of the present invention can make the temperature range of magnetic beads storage wider. Adopt magnetic bead mixed solution of the present invention as nucleic acid (or the carrier particle containing nucleic acid) solvent, can monitor the extraction of nucleic acid simultaneously when nucleic acid is extracted, save the sample adding step of magnetic bead method nucleic acid extraction reagent, can guarantee simultaneously the multiple hole Uniformity of nucleic acid addition concentration. The magnetic bead mixture can stably preserve the nucleic acid.

Description

technical field [0001] The invention relates to the technical field of biochemistry, in particular to a magnetic bead mixture, a nucleic acid preservation method, a nucleic acid extraction reagent and a kit. Background technique [0002] Magnetic bead method nucleic acid extraction reagent is a high-tech product that combines biological science and nanomaterial science. It mainly uses nanotechnology to improve and modify the surface of superparamagnetic nanoparticles to prepare superparamagnetic silicon oxide nanomagnetic particles. beads. The magnetic beads can specifically recognize and efficiently combine with nucleic acid molecules on the microscopic interface. Using the superparamagnetism of silicon oxide nanospheres, under the action of chaotropic salts and an external magnetic field, they can be removed from blood, animal tissues, food, DNA and RNA are isolated from samples such as pathogenic microorganisms, which can be applied in various fields such as clinical dis...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013
Inventor 冼伟杰
Owner FOSHAN UNITED MEDICAL TECH
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