Virus sample direct amplification type preservation solution and application method

A technology for preserving liquids and samples, which is applied in the direction of microorganism-based methods, biochemical equipment and methods, and microorganism measurement/inspection. transport effect

Pending Publication Date: 2021-04-20
苏州白垩纪生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In summary, although the inactivation preservation solution based on guanidinium salt can solve the technical problem of virus inactivation, and at the same time can realize the short-term storage and transportation of RNA in the virus at room temperature, but its components often contain isothiocyanate Guanidine, chelating agents, ethanol or high concentrations of surfactants are common RT-PCR inhibitors, so the subsequent RNA extraction step is still unavoidable

Method used

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  • Virus sample direct amplification type preservation solution and application method
  • Virus sample direct amplification type preservation solution and application method
  • Virus sample direct amplification type preservation solution and application method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0092] Embodiment 1: preservation solution V1

[0093] The present embodiment provides a preservation solution, comprising 0.75% by mass volume of Brij 58, 0.05% lithium dodecyl sulfate, 20 mM trishydroxyaminomethane, 150 mM potassium chloride, 1 mM TCEP, mass volume percentage 0.5% ethanol, 0.05% anthocyanin by mass volume percentage, pH 7.4.

Embodiment 5

[0100] Example 5 Use the preservation solution V1 prepared in Example 1 and the preservation solutions W1-W3 of the comparative example, UTM preservation solution, deionized water, and physiological saline for preservation and direct amplification detection

[0101] With reference to the second aspect of the present invention, the application method of virus sample direct amplification preservation solution is provided, and 10 *5 Pseudovirus (COVID-19-pseudovirus), invert up and down 10 times until mixed, let it stand at room temperature for 10 minutes, then use NEB company Luna universal probe one step RT-qPCR kit (Cat#E3006S), use the method refer to the kit instructions, take 2μL The preservation solution sample containing the pseudovirus was added to the 20 μL system to detect the OFR1ab gene fragment of the pseudovirus. The primer probe sequence was from the CDC in China, and the direct amplification effect of the RNA virus and the presence of the viral RNA were detected a...

Embodiment 6

[0108] Example 6: Verification of different contents of anthocyanins for preservation of viral RNA

[0109] In the formulation of the preservation solution V1 prepared in Example 1, we have prepared a series of preservation solutions containing anthocyanins with different concentrations, and will conduct experiments with reference to the scheme of Example 5. Add 10 *5 Pseudovirus (COVID-19-pseudovirus), invert up and down 10 times until mixed, let it stand at room temperature for 10 minutes, then use NEB company Lunauniversalprobe one step RT-qPCR kit (Cat#E3006S), refer to the kit manual for the usage method, take 2 μL containing The preservation solution samples of the pseudovirus were added to the 20 μL system, and the Ct values ​​of the pseudovirus samples stored in different preservation solutions at room temperature (25° C.) on days 0, 3, 5, and 7 were analyzed by detecting the OFR1ab gene fragment of the pseudovirus. Surprisingly, we found that adding too much anthocya...

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Abstract

The invention belongs to the field of biological sample preservation, and discloses a virus sample direct amplification type sample preservation solution and an application method. The preservation solution comprises a surfactant, a buffer agent, salt ions, a reducing agent, alcohol and a nuclease inhibitor. The preservation solution provided by the invention can quickly inactivate viruses, realizes direct sample loading amplification detection, and avoids a nucleic acid extraction process with tedious steps.

Description

technical field [0001] The invention relates to the field of biological sample preservation. In particular, it relates to a virus sample direct amplification type sample preservation solution and an application method. Background technique [0002] The description of the background technology of the present invention belongs to the relevant technology related to the present invention, and is only used to illustrate and facilitate the understanding of the content of the present invention. prior art on the filing date. [0003] A virus is a special organism without a cellular structure, consisting of a protein shell and genetic material inside. Viruses are tiny and simple in structure, containing only one kind of genetic material (DNA or RNA). Viral infections are extremely harmful, such as SARS or SARS-CoV-2. The initial symptoms of infected patients are mostly fever, fatigue and dry cough, and then gradually develop dyspnea. In severe cases, acute respiratory distress syn...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6806C12R1/93
Inventor 张佳斌邹永龙曲峰何宗顺
Owner 苏州白垩纪生物科技有限公司
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