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Eukaryotic cell nucleic acid extraction method

A cell nucleic acid and extraction method technology, applied in the field of eukaryotic cell nucleic acid extraction, can solve the problems of few types of samples in the extraction method, poor applicability of large-scale extraction, cumbersome extraction steps, etc., and achieve rapid nucleic acid extraction and wide application , the effect of reducing difficulty and complexity

Inactive Publication Date: 2017-01-25
BGI GENOMICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The present invention aims to solve the cumbersome extraction steps, difficult operation, long time-consuming, more reagents required and high cost in the current DNA extraction method, poor applicability of large-scale extraction, and fewer types of samples applicable to the extraction method. question

Method used

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  • Eukaryotic cell nucleic acid extraction method
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  • Eukaryotic cell nucleic acid extraction method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1, establishment of nucleic acid extraction method

[0030] 1. Extraction solution configuration

[0031] The extraction solution is DEB (DNA extraction buffer) solution;

[0032] DEB solution is an alkaline aqueous solution, the solvent is double distilled water, the solute is an alkaline chemical reagent (such as sodium hydroxide or potassium hydroxide), the configuration method is: take a certain amount of double distilled water, add sodium hydroxide or potassium hydroxide powder , stirring continuously until dissolved, and measuring the pH value, the pH value is 8-13, preferably 10.

[0033] 2. Extraction

[0034] Taking the batch extraction of 96-well plate as an example, the specific operation steps are as follows:

[0035] 1) Add the samples to be extracted into 96-well plates;

[0036] 2) Add a certain amount (such as 120 μL) of DEB (DNA extraction buffer) solution (aqueous solution with a pH between 8-13) to 1), and seal the 96-well plate with 8 ...

Embodiment 2

[0039] Nucleic acid extraction in embodiment 2, whole blood

[0040] One, method of the present invention and Chelex-100 extraction method

[0041] Experimental group: extract according to the extract and method in Example 1:

[0042] 1) Add 2 μL of well-mixed isolated human venous whole blood to a 96-well plate;

[0043] 2) Add 120 μL of DEB solution (sodium hydroxide aqueous solution with a concentration of 0.004 g / L; pH value 10) to 1), the volume ratio of whole blood and DEB is 1 / 60, and seal the 96-well plate with a membrane;

[0044] 3) 100°C boiling water bath for 10 minutes; after cooling, get DEB to extract nucleic acid.

[0045] Control group: Chelex-100 extraction method

[0046] 1) Take 10μL of whole blood and add it to a 1.5mL centrifuge tube;

[0047] 2) Add 500 μL of pure water, shake vigorously, and place at room temperature for 15 minutes;

[0048] 3) Centrifuge at 13,000rpm for 3min, remove the supernatant;

[0049] 4) Repeat the above two steps once to...

Embodiment 3

[0073] Example 3, Nucleic Acid Extraction in Dried Blood Sheets

[0074] One, method of the present invention and Chelex-100 extraction method

[0075] test group:

[0076] 1) Add newborn heel blood dried blood slices (obtained by punching) with a diameter of 3mm into a 96-well plate;

[0077] 2) same as 2) of the experimental group of embodiment 2, (concentration is the sodium hydroxide aqueous solution of 0.004g / L; pH value 10,);

[0078] 3) The same as 3) of the experimental group of Example 2, to obtain alkaline DEB to extract nucleic acid.

[0079] Control group:

[0080] 1) Take a dried blood piece with a diameter of 3mm and add it to a 1.5mL centrifuge tube;

[0081] 2)-8) are the same as 2)-8) of the control group in Example 2 to obtain Chelex-100 extracted nucleic acid.

[0082] 2. Comparison of two methods for extracting nucleic acid

[0083] 1. Number of steps and time required for extraction

[0084] Same as embodiment 2;

[0085] 2. Nucleic acid extraction...

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Abstract

The invention discloses a eukaryotic cell nucleic acid extraction method and provides a nucleic acid extraction method which comprises the following steps of: uniformly mixing a to-be-tested sample with alkaline liquid, and performing reaction at a high temperature, thereby obtaining nucleic acid. Experiments prove that compared with an existing DNA extraction method, the method provided by the invention has the advantages that operation steps such as repeated cleaning, warm bath and centrifugation are removed; through creating an alkaline high-temperature liquid environment around the sample, membrane rupture and protein degeneration of eukaryotic cells are accelerated, the extraction efficiency of eukaryon genome DNA is greatly improved, and rapid DNA extraction is realized. The nucleic acid extraction method has no special requirements on aspects of sample types and laboratory conditions, is wide in application range and is suitable for popularization and application.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for extracting eukaryotic nucleic acid. Background technique [0002] Since the advent of PCR technology in the 1980s, its application in molecular biology-related fields has become more and more extensive. Especially in recent years, with the improvement of sensitivity and specificity requirements for clinical diagnosis and the introduction and promotion of the concept of precision medicine, PCR technology has been rapidly and widely used in clinical diagnosis due to its advantages of rapidity, sensitivity and specificity in clinical diagnosis. When it comes to clinical diagnosis and treatment, by detecting the genes in patient samples, it can help doctors diagnose diseases and guide clinical medication. In addition, PCR technology has also been widely used in the fields of judicial identification and agricultural breeding, and it has become a basic technology for the deve...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1003C12Q2527/125
Inventor 郑建超李圳洽袁景风于静张燕王夏曼张红云
Owner BGI GENOMICS CO LTD
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