A kit and method for extracting whole genome dna from blood

A whole-genome and kit technology, which is applied to the whole-genome DNA extraction kit in blood and the field thereof, to achieve the effects of improving accuracy, improving sensitivity and reducing interference

Active Publication Date: 2017-01-11
ZICHENG RUISHENGHUI BEIJING BIOTECH DEV CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The invention provides a kit for efficiently extracting whole-genome DNA from blood and a method for extracting whole-genome DNA from blood, so as to solve the problems of reducing the impact of organic solvents or strong alkalis on the health of operators and subsequent experiments, and the like. Efficient extraction of high-purity whole-genome DNA even from trace amounts of blood samples

Method used

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  • A kit and method for extracting whole genome dna from blood
  • A kit and method for extracting whole genome dna from blood

Examples

Experimental program
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Effect test

Embodiment 1

[0084] Example 1 Kit for extraction of whole genome DNA from blood

[0085] The components of the kit are as follows:

[0086] 1) Red blood cell lysate: deionized ultrapure water (ddH 2 O), the resistivity is greater than 18MΩ*cm), sterilize for 20 minutes at 121°C and 100 kPa pressure; after sterilization, each 400ml is packed into 500ml sterile plastic bottles;

[0087] 2) White blood cell washing solution: add 50ml Triton-X100 to 400ml sterile deionized ultrapure water, and dilute to 500ml at 68°C to prepare a 10% Triton-X100 mother liquor; then add 10ml 10% Triton-X100 mother liquor Add 990ml sterile deionized ultrapure water to prepare a 0.1% Triton-X100 leukocyte washing solution; each 300ml is divided into 300ml sterile plastic bottles;

[0088] 3) Digestive juice: 60.55g Tris alkali dissolved in 400ml ddH 2 In O, adjust the pH value to 8.0 with HCl (add about 2.0ml for concentrated HCl), use ddH 2 O dilute to 500ml, sterilize at 121℃, 100 kPa pressure for 20 minutes to prepare...

Embodiment 2

[0101] Example 2 Using the kit of the present invention to extract whole genome DNA from cryopreserved blood

[0102] Collect 8 clinical diabetic retinopathy patients and 7 normal people with 5ml EDTA anticoagulant blood (a scientific research cooperation project with a partner unit, approved by the hospital ethics committee, and signed informed consent with volunteers).

[0103] 1) Lysis of blood red blood cells: first remove the frozen blood from the -80°C refrigerator and put it in a 37°C water bath until the blood sample melts. Add 8ml of erythrocyte lysate to each 15ml centrifuge tube, and add 1ml of blood sample, shake and mix by hand, place at 4℃ for 20min and centrifuge at 2000rpm for 15min at low speed; after centrifugation, discard the supernatant;

[0104] 2) White blood cell washing: Add 2ml of white blood cell washing solution to a 15ml centrifuge tube containing white blood cell pellets, mix well, put it in a low-speed horizontal centrifuge, centrifuge at 2000 rpm for 1...

Embodiment 3

[0111] Example 3 Using the kit of the present invention to extract whole genome DNA from blood stains

[0112] Take 1ml of anticoagulant blood from 8 clinical patients and add them to worn-out clothes and leave them at room temperature for 2-3 days. Cut the cloth stained with blood, put them into a 50ml centrifuge tube with 30ml red blood cell lysate, vortex for 5min, place at 4℃ for 20min, take out the cloth with tweezers, let the liquid drip as much as possible, and centrifuge at 2000rpm at low speed 15min; discard the supernatant after centrifugation;

[0113] Add 30ml of white blood cell washing solution to a 50ml centrifuge tube containing white blood cell pellet, mix well, centrifuge at low speed at 1000rpm for 10 minutes, discard the supernatant carefully, and repeat the washing twice; the rest of the steps follow step 3 in Example 2 -7 is completed, the extracted whole genome DNA is detected by 0.8% agarose gel electrophoresis, Figure II It shows that the whole genomic DN...

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Abstract

The invention relates to a kit of extracting a whole genome DNA (Deoxyribonucleic Acid) from blood and a using method thereof. The kit is characterized by comprising a red blood cell lysate, a white blood cell scrubbing solution, digestive juice, proteinase K, a purifying liquid, gDNA salting out liquid, a gDNA scrubbing solution, a gDNA eluant and the like. The using method of the whole genome DNA extraction kit for blood is characterized by comprising the following steps: washing the red blood cell split to obtain the white blood cell; splitting the white blood cell by the digestive juice containing the proteinase K; and further purifying by an improved lithium chloride purifying liquid, salting out the liquid layer, and carrying out chromatography to obtain the high purity whole genome DNA. When the kit provided by the invention is used to extract the whole genome DNA in blood, plasma and serum in blood are not separated in advance but fresh or frozen anti-freezing whole blood is taken, wherein the lowest blood volume required reaches 20 microliters or blood cakes are required. According t the kit provided by the invention, the whole genome DNA with high purity can be fully unlinked and the PCR (Polymerase Chain Reaction) amplification is efficiently carried out, so that the kit is used for scientific research or clinical diagnostic analysis such as PCR amplification, gene expression, gene sequencing, whole genome sequencing, exome sequencing, gene mutation and single nucleotide polymorphism.

Description

Technical field [0001] The invention relates to a kit for extracting whole genome DNA from blood and a method for extracting whole genome DNA from blood by using the kit. When the kit of the present invention extracts whole genome DNA from blood, there is no need to separate plasma and serum from blood in advance, only fresh or frozen anticoagulated whole blood is taken, and the minimum required blood volume can reach 20 μl or blood spots. . The high-purity whole genome DNA obtained by its efficient extraction can be used for scientific research or clinical diagnostic analysis such as PCR amplification, gene expression, gene sequencing, whole genome sequencing, exome sequencing, gene mutation and single nucleotide polymorphism . . Background technique [0002] Genomic DNA (gDNA) is all genetic information including coding and non-coding sequences in haploid cells. All life information can be found from the whole genome DNA, including the problems of gene expression, gene muta...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/10
Inventor 曾沃坦黄启明梁辰陈建勋
Owner ZICHENG RUISHENGHUI BEIJING BIOTECH DEV CO LTD
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