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A method for fast nucleic acid extraction

An extraction method and nucleic acid technology, applied in biochemical equipment and methods, DNA preparation, microbial measurement/inspection, etc., can solve the problems of complex extraction process and long time-consuming

Inactive Publication Date: 2019-01-15
博迪泰(厦门)生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the invention is to provide a rapid nucleic acid extraction method, which solves the problems of complicated extraction process, long time-consuming and special equipment

Method used

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  • A method for fast nucleic acid extraction
  • A method for fast nucleic acid extraction
  • A method for fast nucleic acid extraction

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] PCR results for the detection of Mycoplasma pneumoniae in nasopharyngeal swab samples from DNA lysates. Proceed as follows:

[0039] 1. Dip the nasopharyngeal swab with the sample directly into 1ml DNA lysis solution, 1ml DNA lysis solution is 1.5M guanidine hydrochloride, 50mM Tris (PH=8.0), 100mM NaCl, 5mM EDTA and 1% Tween20, and treat for 10 seconds .

[0040] 2. Immerse the cellulose filter paper in the DNA lysate in step 1 for 10 seconds.

[0041] 3. Immerse the cellulose filter paper obtained in the step 2 into the rinsing liquid, the rinsing liquid is 10 mM Tris pH=8.0 and 0.1% Tween-20, and treat for 10 seconds.

[0042] 4. Immerse the cellulose filter paper obtained in step 3 into the PCR reaction system for 10 seconds, or leave it in the reaction system.

[0043] 5. PCR was amplified for 90 minutes. The same sample extracted from the standard plasmid and bacterial genomic DNA extraction kit (Beijing Tiangen) was used as a positive control, and the nasophar...

Embodiment 2

[0046] Detection of RPA results for influenza A in nasopharyngeal swab samples treated with RNA lysate. Proceed as follows:

[0047] 6. Immerse the nasopharyngeal swab with the sample directly into 1ml RNA lysis solution, 1ml RNA lysis solution is 4.0M guanidine isothiocyanate, 25mM sodium citrate, 0.5% (m / V) sodium lauryl creatine, 0.1mM β Mercaptoethanol, process for 10 seconds.

[0048] 7. Soak the cellulose filter paper in the RNA lysate obtained in step 6 for 10 seconds.

[0049] 8. Immerse the cellulose filter paper obtained in step 7 in the rinse solution, which is 10 mM Tris (pH=8.0) and 0.1% Tween-20, for 10 seconds.

[0050] 9. Immerse the cellulose filter paper obtained in step 8 into the RPA reaction system for 10 seconds, or leave it in the reaction system.

[0051] 10. RPA was amplified for 15 minutes. The same sample extracted from the standard virus strain and the virus RNA extraction kit (Shanghai Sangong) was used as a positive control, and the nasopharyng...

Embodiment 3

[0054] Detection of RPA results for dengue fever in RNA lysate-treated serum samples. Proceed as follows:

[0055] 11. Add 300ul of dengue fever patient serum sample to 1.2ml RNA lysate, the RNA lyse solution is 50μg / ml Proteinase K, 25mM sodium citrate, 0.5% (m / V) sodium laurate, 4.0M guanidine isothiocyanate and 0.1 mM β-mercaptoethanol,

[0056] 12. Treat with 50 μg / ml Proteinase K, 25 mM sodium citrate, 0.5% m / V sodium laurel creatine and 0.1 mM β-mercaptoethanol for 1 hour, then add 4.0 M guanidine isothiocyanate for 10 seconds.

[0057] 13. Soak the cellulose filter paper in the RNA lysate obtained in step 12 for 10 seconds.

[0058] 14. Immerse the cellulose filter paper obtained in step 13 into a rinsing buffer, and the rinsing solution is 10 mM Tris (pH=8.0) and 0.1% Tween-20 for 10 seconds.

[0059] 15. Immerse the cellulose filter paper obtained in step 14 into the RPA reaction system for 10 seconds, or leave it in the reaction system.

[0060] 16. RPA was ampli...

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Abstract

A method for extract nucleic acid includes such step as cracking nucleic acid sample, adding sample containing nucleic acid to cracking solution to form nucleic acid cracking solution, cracking nucleic acid sample to form nucleic acid cracking solution, cracking nucleic acid sample to obtain nucleic acid sample, cracking nucleic acid sample to obtain nucleic acid sample, cracking nucleic acid sample to obtain nucleic acid sample, cracking nucleic acid sample to obtain nucleic acid sample, cracking nucleic acid sample to obtain nucleic acid sample, cracking nucleic acid sample to obtain nucleicacid sample. 2, adsorb nucleic acid: adding that carry into the nucleic acid cleavage solution to obtain a carrier for adsorbing nucleic acid; 3, remove impurities: rin that carrier adsorbed with nucleic acid in a rinsing solution to remove impurities other than nucleic acid; 4, amplification reaction, wherein that rinse carrier adsorb with nucleic acid is directly added into a nucleic acid amplification system for amplification reaction. The carrier is cellulose filter paper; a simple, a method for extract nucleic acid based on cellulose filt paper is disclosed. Applicable to polymerase chain reaction amplification and recombinase polymerase isothermal amplification and other nucleic acid amplification technology, suitable for operating at room temperature, suitable for biological samples from various sources, can be loaded with nucleic acid cellulose filter paper directly into the nucleic acid amplification system for amplification reaction, the whole process is less time-consuming.

Description

technical field [0001] The invention relates to the field of nucleic acid extraction, in particular to a rapid nucleic acid extraction method. Background technique [0002] The main challenge for molecular diagnosis using nucleic acid amplification technology is the pretreatment of nucleic acid-containing materials before amplification and the complicated nucleic acid extraction steps. The currently widely used silica- and magnetic-bead-based techniques suffer from a number of drawbacks, including low efficiencies (≤10%); and the need to optimize for specific target types, resulting in extraction procedures for one sample type often being different from others. The sample types are incompatible; while the method utilizing phenol-chloroform has a high recovery rate, its scope of application is limited due to the high toxicity of the extract components; the method based on detergent has a simple dissolution process, but requires high-temperature denaturation (95 ℃) treatment,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1003C12Q2521/537
Inventor 李博安张睿
Owner 博迪泰(厦门)生物科技有限公司
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