Kit and application thereof

A kit and cell nucleus technology, applied in the biological field, can solve problems that need to be improved, and achieve the effects of high-throughput automated nucleic acid extraction, strong lysis, and accelerated DNA release

Active Publication Date: 2017-05-31
TIANGEN BIOTECH BEIJING
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Currently, existing genomic DNA ex

Method used

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  • Kit and application thereof
  • Kit and application thereof
  • Kit and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1 Contrast experiment of manual operation extracting blood

[0039] Using this method to extract 2ml of blood samples, and Tiangen Biochemical Technology (Beijing) Co., Ltd. product DP333 a large number of blood genomic DNA extraction kits for comparison experiments with magnetic bead method and spin column method extraction. DP333 is operated according to the existing manual. The magnetic bead method extraction steps are as follows:

[0040] (1) Blood pretreatment:

[0041] a. Place 5ml of blood samples No. 1-3 at room temperature until completely thawed, centrifuge at 2500rpm for 10 minutes in a centrifuge, discard the upper part of the plasma, and keep the middle and lower part of the blood cells.

[0042] b. Add an equal volume of 2.5ml of nucleus separation solution and 25mg of glass beads into a blood-drawing tube containing 2.5ml of blood cells, invert and mix 5 times, and centrifuge at 3600rpm for 10min.

[0043]c Take out the blood vessel and deca...

Embodiment 2

[0059] Example 2 Genomic DNA extraction of 10ml blood on Thermo KingFisher Flex magnetic rod method automation instrument

[0060] 1. Reagents and materials:

[0061] The nuclei isolation solution contains:

[0062] 30mmol / L Tris;

[0063] 0.015% Triton X-100;

[0064] 5mmol / L EDTA;

[0065] 30mmol / L MgCl 2 ;as well as

[0066] 30mmol / L KCl, the pH value of the cell nucleus separation solution is 7.5.

[0067] The nuclear lysate contains:

[0068] 4.5mol / L guanidine hydrochloride;

[0069] 1.5mol / L guanidine isothiohydrogen;

[0070] 40mmol / L Tris (pH=8.0);

[0071] 40mmol / L EDTA;

[0072] 4% Triton X-100; and

[0073] 0.3% sodium deoxycholate, the pH of the cell nucleus lysate is 8.0.

[0074] Glass beads were purchased from Suzhou Jinxing Glass Abrasive Co., Ltd., and the diameter of the glass beads was 1 mm.

[0075] Deproteinization buffer contains:

[0076] 5mol / L guanidine hydrochloride;

[0077] 40mmol / L Tris (pH=8.0);

[0078] 40mmol / L EDTA;

[0079] 1...

Embodiment 3

[0113] Example 3 Genomic DNA extraction of 2ml of blood was carried out on the automatic instrument of Boao Bio Labkeeper pipetting method

[0114] 1. Materials and reagents

[0115] The nuclei isolation solution contains:

[0116] 30mmol / L Tris;

[0117] 0.015% Triton X-100;

[0118] 5mmol / L EDTA;

[0119] 30mmol / L MgCl 2 ;as well as

[0120] 30mmol / L KCl, the pH value of the cell nucleus separation solution is 7.5.

[0121] The nuclear lysate contains:

[0122] 4.5mol / L guanidine hydrochloride;

[0123] 1.5mol / L guanidine isothiohydrogen;

[0124] 40mmol / L Tris (pH=8.0);

[0125] 40mmol / L EDTA;

[0126] 4% Triton X-100; and

[0127] 0.3% sodium deoxycholate, the pH of the cell nucleus lysate is 8.0.

[0128] Glass beads, the diameter of the glass beads is 0.5 mm.

[0129] Deproteinization buffer contains:

[0130] 5mol / L guanidine hydrochloride;

[0131] 40mmol / L Tris (pH=8.0);

[0132] 40mmol / L EDTA;

[0133] 1.5% Sodium Lauroyl Sarcosinate; and

[0134] 30%...

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Abstract

The invention relates to a kit and application thereof. Particularly, the kit comprises a cell nucleus separating solution, wherein the cell nucleus separating solution is prepared from 20 to 50 mmol/L of Tris, 0.01 to 0.2 percent of Triton X-100, 1 to 10 mmol/L of EDTA, 5 to 50 mmol/L of MgCl2, and 5 to 50 mmol/L of KCl. Therefore, the kit can be used for effectively pretreating blood, so as to obtain cell nucleus sediments for subsequent treatment.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a kit for extracting blood genome DNA and its application. Background technique [0002] The extraction of blood genomic DNA is the first step in molecular medical testing, and the acquisition of high-quality DNA is the key to ensuring the success of subsequent testing. For genetic disease-related gene detection and research on methylation sites, more detection sites are required, and the demand for genomic DNA is greater, so many researchers need to extract from larger volumes of blood, such as 1-10ml Genomic DNA to meet subsequent needs. [0003] Currently, existing genomic DNA extraction methods still need to be improved. Contents of the invention [0004] The present invention has been achieved by the inventors based on the following findings and facts: [0005] The inventors have found that the currently commonly used genomic DNA extraction methods have obvious deficiencies...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1013C12Q2527/119C12Q2527/125C12Q2563/143C12Q2563/149
Inventor 邹爱兰刘玉方李晓晨孙克非
Owner TIANGEN BIOTECH BEIJING
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