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Virus nucleic acid extraction alcohol-free lysis buffer, kit and extraction method

A virus nucleic acid and lysate technology, applied in the field of biomedicine, can solve problems such as potential safety hazards, and achieve the effects of consistent accuracy, equivalent linear range, and simple preparation methods

Active Publication Date: 2022-03-11
GENFINE BIOTECH BEIJING CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are safety hazards for both the preparation solution and the people who use it

Method used

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  • Virus nucleic acid extraction alcohol-free lysis buffer, kit and extraction method
  • Virus nucleic acid extraction alcohol-free lysis buffer, kit and extraction method
  • Virus nucleic acid extraction alcohol-free lysis buffer, kit and extraction method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Embodiment 1 Determination of surfactant composition

[0084] (1) Comparison of different surfactants used alone

[0085] The lysates used separately with different surfactants were prepared, and the compositions of different lysates are shown in Table 1.

[0086] Table 1 The lysate composition of different surfactants used alone

[0087]

[0088] The experimental process is as follows: the extraction reagent uses the "Virus Nucleic Acid Extraction Kit" (Y502-G10) produced by Jifan Biological Company, which includes magnetic bead preservation solution, magnetic bead lysis solution, washing solution, rinsing solution and eluent . The detection kit uses Harbin Guosheng Biology "African Swine Fever Virus Fluorescent PCR Detection Kit B Box", which includes fluorescent PCR reaction system, sterile nuclease-free water, primer-probe mixture, negative control, and positive control. As the quality control product, ASF pseudovirus quality control product (Concentration of ...

Embodiment 2

[0114] Embodiment 2 Determination of the concentration of guanidinium salt

[0115] Nucleic acid was extracted according to the steps in Example 1, and PCR amplification was performed on the extracted nucleic acid. Only the lysate was replaced with the lysate in Table 11. The experimental scheme is shown in Table 12, and the experimental results are shown in Table 13.

[0116] Table 11 The lysates of different guanidinium salt concentrations

[0117]

[0118] Table 12 Experimental scheme of Example 2

[0119]

[0120] Table 13 Extraction of 10,000 copies of African swine fever quality control products from the experimental results of Example 2

[0121]

[0122] Analysis of results: The effect of guanidinium salt concentration in the range of 3-4M basically has little difference, so follow-up research uses 3M guanidine isothiocyanate to continue the research.

Embodiment 3

[0123] Example 3 Determination of buffer pH value

[0124] Nucleic acid was extracted according to the steps in Example 1, and PCR amplification was performed on the extracted nucleic acid, only the lysate was replaced with the lysate in Table 14, the experimental scheme was shown in Table 15, and the experimental results were shown in Table 16.

[0125] Table 14 The lysate composition of different buffer pH values

[0126]

[0127] Table 15 Experimental scheme of Example 3

[0128]

[0129] Table 16 Extraction of 10,000 copies of African swine fever quality control products from the experimental results of Example 3

[0130]

[0131] Result analysis: The pH range of the buffer solution is within the range of pH7.5-8.5, and the extraction performance is stable and good, and the pH7.5 is selected for further research.

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Abstract

The invention discloses a virus nucleic acid extraction alcohol-free lysis solution, a kit and an extraction method. The alcohol-free lysis solution for extracting viral nucleic acid consists of guanidine salt, inorganic salt, a surfactant and a buffer solution, the guanidine salt is any one or two of guanidine isothiocyanate and guanidine hydrochloride; the inorganic salt is any one or two of sodium chloride and potassium chloride; the surfactant is polyethylene glycol and Tween 20; the pH value of the buffer solution is 7.5 to 8.5. According to the invention, the harm of alcohol volatilization or pungent smell in the traditional nucleic acid extraction lysate to human bodies can be effectively avoided; the preparation method is simple, free of toxic chemical reagents, safe and pollution-free, and can be used for manual extraction and also can be used for an automatic platform; compared with an alcohol lysis solution, the virus nucleic acid detection sensitivity is equivalent, the accuracy is consistent, and the linear range is equivalent.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to an alcohol-free lysate for extracting viral nucleic acid, a kit and an extraction method. Background technique [0002] Nucleic acid is a biological macromolecular compound synthesized by many nucleotides, and is one of the most basic substances of life. Nucleic acid extraction is the most basic method to obtain nucleic acid. There are many methods for nucleic acid extraction, such as solution method, filter column method, and other methods such as magnetic bead method that have developed rapidly in recent years. [0003] Magnetic bead nucleic acid extraction has incomparable advantages over traditional DNA extraction methods, mainly reflected in: 1. It can realize automation and large-scale operations. At present, there are 96-channel nucleic acid automatic extraction instruments, which can be achieved in the extraction time of one sample. The processing of 96 s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013Y02A50/30
Inventor 王鹏韩典霖俞萍
Owner GENFINE BIOTECH BEIJING CO LTD
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