Kit and method for extracting blood genome DNA based on magnetic bead technology

A kit and genome technology, applied in the field of molecular biology, can solve the problems of low purity, insufficient white blood cell lysis, affecting PCR amplification efficiency, etc., and achieve the effects of reducing nucleic acid loss, shortening extraction time, and increasing extraction efficiency.

Pending Publication Date: 2018-05-25
HANGZHOU LC BIOTECH
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  • Claims
  • Application Information

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Problems solved by technology

This method avoids the use of toxic organic solvents, and the extracted DNA has a high purity, but it requires a large amount of blood samples, and it takes a long time to digest the white blood cells with protease alone. In addition, the use of strong alkali is not good for the health of the operator. It will al

Method used

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  • Kit and method for extracting blood genome DNA based on magnetic bead technology
  • Kit and method for extracting blood genome DNA based on magnetic bead technology
  • Kit and method for extracting blood genome DNA based on magnetic bead technology

Examples

Experimental program
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Effect test

Embodiment 1

[0028] Preparation of reagents in whole blood genomic DNA extraction kit.

[0029] (1) Lysis solution: a mixed solution composed of 50 mM Tris-HCl, 10 mM EDTA, 0.5 M NaCl and 1% SDS by volume. Specific preparation method: Weigh 7.88g Tris-HCl, 2.92g EDTA and 29.22g NaCl, add 800mL deionized water and stir to dissolve, add 10mL SDS, continue to add deionized water to make up to 1L.

[0030] (2) Red blood cell lysate: 100mM NH 4 Cl, 10mM KHCO 3 and 0.1 mM Na 2 Mixed solution of EDTA. Specific preparation method: weigh 5.34g NH 4 Cl, 1.0g KHCO 3 and 0.03g Na 2 EDTA, add deionized water, stir evenly, and dilute to 1L.

[0031] (3) Magnetic bead rinsing solution: ethanol solution with a volume fraction of 70%. Specific preparation method: Measure 700mL of absolute ethanol and add 300mL of deionized water, mix well.

[0032] (4) DNA eluent: 10 mM Tris-HCl solution, pH 8.0. Specific preparation method: Weigh 1.58g Tris-HCl and add deionized water to make the volume to 1L, a...

Embodiment 2

[0034] Genomic DNA was extracted from 10 human frozen anticoagulated blood using the kit in Example 1.

[0035] ①Take 200 μL of thawed blood sample, add 400 μL of erythrocyte lysate, mix well, let stand at room temperature for 1 min, centrifuge at 12000 g for 2 min, and discard the supernatant.

[0036] ② Add 100 μL of lysate to the pellet, mix well and incubate at 70°C for 10 minutes.

[0037] ③ Add 100 μL of magnetic bead solution, mix well, let stand at room temperature for 5 minutes, place on a magnetic stand for 5 minutes, and discard the supernatant.

[0038] ④ Add 200 μL magnetic bead washing solution to resuspend the magnetic bead nucleic acid mixture, wash twice, and blot the residual liquid.

[0039] ⑤ Open the lid and let stand at room temperature for 5 minutes, add 50 μL of DNA eluent, mix well by pipetting, and let stand at room temperature for 2 minutes.

[0040] ⑥Place it on the magnetic stand for 5 minutes, transfer the supernatant to a new nuclease-free tube...

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Abstract

The invention discloses a kit and method for extracting blood genome DNA based on a magnetic bead technology, and belongs to the field of molecular biology. The kit comprises five components, namely lysate, red blood cell lysate, magnetic beads, a magnetic bead bleaching solution and a DNA eluent. The kit disclosed by the invention does not contain an organic solvent; after complete blood cells are split, DNA molecules are adsorbed through the magnetic beads; impurities are removed through bleaching; finally, the DNA molecules on the magnetic beads are eluted, so that the high-quality genome DNA is obtained. The kit has the advantages of simple and quick operation, high extraction efficiency, low cost and the like. The extracted blood genome DNA is high in purity, and can accord with subsequent scientific researches such as PCR amplification and gene sequencing or clinical diagnosis and analysis.

Description

technical field [0001] The invention belongs to the field of molecular biology, and relates to a kit for extracting blood genome DNA based on magnetic bead technology, which can be applied to extract DNA from fresh or frozen anticoagulated blood of different species. The invention also relates to a method for extracting DNA from blood samples using the kit. Background technique [0002] Blood is an opaque red liquid that flows in the heart and blood vessels, and its main components are plasma and blood cells. Genomic DNA (genomic DNA) is the carrier of genetic information, which is all genetic information in haploid cells, including coding sequences and non-coding sequences. Extracting high-quality DNA from human or animal blood is an important prerequisite for genetic testing, precision medicine and other molecular biology research. The general procedures for blood DNA extraction and purification are: break red blood cells, centrifuge white blood cells, break white blood ...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1013
Inventor 郎秋蕾梁洪殷楠楠袁丹娄奇彬李倩
Owner HANGZHOU LC BIOTECH
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