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Nucleic acid purifying reagent and method

A reagent and nucleic acid technology, applied in the field of nucleic acid purification reagents, can solve the problems of reducing the number of disposable commercial kits used, nucleic acid recovery rate and unsatisfactory purity, and achieve the effect of improving purification effect, improving purification effect and improving purification efficiency.

Active Publication Date: 2020-06-05
INTEGRATED BIOSYSTEMS CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The PCR purification reagent of the invention can be successfully used for the purification of DNA without affecting the yield of DNA purification. The use of the PCR purification reagent together with the regenerated silicon column can reduce the number of disposable commercial kits used, reduce the generation of laboratory waste, and protect Environment, save social resources and use costs, but the recovery rate and purity of nucleic acid are not ideal for this application

Method used

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  • Nucleic acid purifying reagent and method
  • Nucleic acid purifying reagent and method
  • Nucleic acid purifying reagent and method

Examples

Experimental program
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Effect test

Embodiment 1

[0075] A nucleic acid purification reagent:

[0076] Silicone hydroxyl magnetic beads with a particle size of 300nm at a concentration of 20mg / mL;

[0077] DNA binding solution: 4M guanidine hydrochloride, 0.35M sodium acetate pH=4.5, 1% Triton X-100;

[0078] Washing solution: Reagent 1: 4M guanidine hydrochloride, 0.25M sodium acetate pH=4.5, 1% Triton X-100;

[0079] Reagent 2: ethanol with a volume fraction of 70%, 0.1M sodium acetate pH=4.5, 10mM sodium citrate pH=4.5;

[0080] Eluent: 5mM Tris-HCl pH=8.

[0081] Purification method:

[0082] (1) Magnetic bead pretreatment: Shake 5 μL of silanol magnetic bead particles evenly, suck them into a centrifuge tube, perform magnetic separation, discard the supernatant, and obtain pretreated silanol magnetic bead particles;

[0083] (2) Mixing: Add 70 μL of DNA binding solution and 15 μL of sample to the pretreated silanol magnetic bead particles obtained in step (1), ultrasonically mix and let stand for 90 s, magnetically s...

Embodiment 2

[0089] A nucleic acid purification reagent:

[0090] Silicone hydroxyl magnetic beads with a particle size of 800nm ​​at a concentration of 30mg / mL;

[0091] DNA binding solution: 8M guanidine hydrochloride, 0.2M sodium cocoyl glutamate, 0.4M sodium lauroyl glutamate, 0.45M sodium acetate pH=5, 3% Triton X-100;

[0092] Washing solution: Reagent 1: 8M guanidine hydrochloride, 0.35M sodium acetate pH=5, 3% Triton X-100;

[0093] Reagent 2: ethanol with a volume fraction of 70%, 0.2M sodium acetate pH=5, 20mM sodium citrate pH=5;

[0094] Eluent: 15 mM Tris-HCl pH=9.

[0095] Purification method:

[0096] (1) Magnetic bead pretreatment: Shake 15 μL silanol magnetic bead particles evenly, suck them into a centrifuge tube, perform magnetic separation, discard the supernatant, and obtain pretreated silanol magnetic bead particles;

[0097] (2) Mixing: pipette 90 μL of DNA binding solution and 25 μL of sample into the pretreated silanol magnetic bead particles obtained in step (...

Embodiment 3

[0103] A nucleic acid purification reagent:

[0104] Silanol magnetic beads with a particle size of 500nm at a concentration of 25mg / mL;

[0105] DNA binding solution: 6M guanidine hydrochloride, 0.2M sodium cocoyl glutamate, 0.6M sodium lauroyl glutamate, 0.40M sodium acetate pH=4.7, 2% Triton X-100;

[0106] Washing solution: Reagent 1: 6M guanidine hydrochloride, 0.30M sodium acetate pH=4.7, 2% Triton X-100;

[0107] Reagent 2: ethanol with a volume fraction of 70%, 0.15M sodium acetate pH=4.7, 15mM sodium citrate pH=4.6;

[0108] Eluent: 10 mM Tris-HCl pH=8.5.

[0109] Purification method:

[0110] (1) Magnetic bead pretreatment: Shake 10 μL of silanol magnetic bead particles evenly, suck them into a centrifuge tube, perform magnetic separation, discard the supernatant, and obtain pretreated silanol magnetic bead particles;

[0111] (2) Mixing: Add 80 μL of DNA binding solution and 20 μL of sample to the pretreated silanol magnetic bead particles obtained in step (1), ...

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Abstract

The invention discloses a nucleic acid purifying reagent and method, and belongs to the biological field. The reagent contains magnetic beads, DNA binding liquid, a scrubbing solution and eluant, wherein the magnetic beads are silicon-hydroxyl-modified magnetic beads; the DNA binding liquid is guanidine hydrochloride, sodium acetate and Triton X-100; the scrubbing solution includes a reagent 1 including guanidine hydrochloride, sodium acetate and Triton X-100 as well as a reagent 2 including ethanol, sodium acetate and sodium citrate; and the eluant is Tris-HCl. By adding sodium cocoyl glutamate and sodium lauryl glutamate into the DNA binding liquid in a concentration ratio of 1 to (1-5) in an implementation process, a purification effect can be obviously improved. By virtue of interaction of the reagents, the purification efficiency of nucleic acid is obviously improved, the purity is high, and the separation purification yield can reach up to 98.76% and is obviously higher than thatin the prior art.

Description

technical field [0001] The invention belongs to the biological field, and in particular relates to a nucleic acid purification reagent and a nucleic acid purification method using the reagent. Background technique [0002] With the advancement of technology, nucleic acid purification has become an essential work in the preparation stage of molecular biology research. The first step in experimental operations such as disease diagnosis, cloning, sequencing, amplification, hybridization, and cDNA analysis is to purify complete and high-quality nucleic acids, otherwise a large number of intracellular components (such as proteins, polysaccharides) or certain components in the lysate The presence of components (organic solvents, metal ions) can hinder the completion of these molecular biological downstream reactions. [0003] Nucleic acid includes deoxyribonucleic acid and ribonucleic acid, and most of them exist in the state of binding to proteins in cells. The general principl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013
Inventor 张瑜高静蔡亦梅范东雨任鲁风
Owner INTEGRATED BIOSYSTEMS CO LTD
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