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RNA protective agent and application thereof

A technology of protective agent and chaotropic agent, which is applied in the direction of biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problem of reducing the difficulty and cost of extraction, the difficulty of extracting large volumes of urine samples, and increasing the extraction of original urine samples Volume and other issues

Pending Publication Date: 2022-04-29
HUNAN YEARTH BIOTECHNOLOGICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The urine sample preserved by the preservation solution of the present invention can also enrich the precipitate by low-speed centrifugation, and then extract the RNA in the precipitate by the conventional Trizol method, which provides a convenient way for downstream extraction and increases the extraction volume of the original urine sample , which solves the problem of difficult extraction of large volumes of urine samples, and greatly reduces the difficulty and cost of extraction

Method used

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  • RNA protective agent and application thereof
  • RNA protective agent and application thereof
  • RNA protective agent and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Embodiment 1: the synergy of chaotropic agent and sedimentation aid in this preservation solution

[0051] Collect morning urine samples from normal people. After collection, take 16ml of urine and add them to 50ml centrifuge tubes / purified water centrifuge tubes containing the following groups of preservation solutions. The volume ratio of purified water or preservation solution to urine is 1:4. The room temperature is 23.6°C, and the solvent is sterilized pure water.

[0052] Table 1

[0053]

[0054]

[0055] The urine of the first group of purified water was immediately enriched and precipitated, and the precipitate was added with the RZ reagent in the Tiangen total RNA extraction kit (DP419) and stored in a -80°C refrigerator for later use. After 5 days, total urine RNA was extracted. The urine of group 2 purified water was collected and placed at room temperature for 5 days, then enriched and precipitated with other preservation solution groups, and then e...

Embodiment 2

[0061] Example 2: Nuclease Inhibitor Test

[0062] Collect morning urine samples from normal people. After collection, take 16ml of urine and add them to 50ml centrifuge tubes / purified water centrifuge tubes containing the following groups of preservation solutions. The volume ratio of purified water or preservation solution to urine is 1:4. The room temperature is 23.4°C, and the solvent is sterilized pure water.

[0063] table 3

[0064] Group No components 1 Purified water, enriched sediment at 0 days 2 Purified water, placed at room temperature for 5 days 3 4M Guanidine HCl, 20% PEG, 50mM EDTA 4 4M Guanidine Isothiocyanate, 20% PEG, 50mM EDTA 5 4M Guanidine HCl, 20% PEG, 50mM EGTA 6 4M Guanidine Isothiocyanate, 20% PEG, 50mM EGTA 7 4M Guanidine Hydrochloride, 20% PEG, 100mM Glyceraldehyde 8 4M Guanidine Isothiocyanate, 20% PEG, 100mM Glyceraldehyde 9 4M Guanidine Hydrochloride, 20% PEG, 5mM ATA 10 4M ...

Embodiment 3

[0070] Embodiment 3: reducing agent test

[0071] On the basis of group 3 (4M guanidine hydrochloride, 20% PEG, 50mM EDTA) and group 4 (4M guanidine isothiocyanate, 20% PEG, 50mM EDTA) of embodiment 2, it is expected to improve the RNA acquisition rate by adding a reducing agent .

[0072] Collect morning urine samples from normal people. After collection, take 16ml of urine and add them to 50ml centrifuge tubes / purified water centrifuge tubes containing the following groups of preservation solutions. The volume ratio of purified water or preservation solution to urine is 1:4. The room temperature is 24.1°C, and the solvent is sterilized pure water.

[0073] table 5

[0074] Group No components 1 Purified water, enriched sediment at 0 days 2 Purified water, placed at room temperature for 5 days 3 4M Guanidine HCl, 20% PEG, 50mM EDTA 4 4M Guanidine Isothiocyanate, 20% PEG, 50mM EDTA 5 4M Guanidine Hydrochloride, 20% PEG, 50mM EDTA, 8...

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Abstract

The invention discloses an RNA (Ribonucleic Acid) protective agent and application thereof. The protective agent comprises the following components: a chaotropic agent, a settling agent, a reducing agent, a nuclease inhibitor, a surfactant, lower alcohol, a buffer solution and a solvent. Wherein the chaotropic agent is guanidine isothiocyanate or guanidine hydrochloride; the reducing agent is tris (2-carboxyethyl) phosphine hydrochloride; the settling aid is at least one of PEG (Polyethylene Glycol) and sulfosalicylic acid. The protective agent can effectively maintain the stability of RNA in the transported and preserved urine sample, more importantly, the stability of RNA in the normal-temperature transportation process can be guaranteed, and the protective agent is suitable for preservation of various complex urine samples clinically. The urine sample preserved by the preserving fluid can be enriched and precipitated in a low-speed centrifugation manner, and RNA in the precipitate is extracted by a conventional Trizol method, so that a convenient manner is provided for downstream extraction, the extraction volume of the original urine sample is increased, the problem that the large-volume urine sample is difficult to extract is solved, and the extraction efficiency of the urine sample is improved. The extraction difficulty and the extraction cost are greatly reduced, and the method has a very good application prospect.

Description

technical field [0001] The invention belongs to the technical field of biological RNA sample preservation, and in particular relates to a composition protectant for storing RNA in liquid samples at room temperature and an application method thereof. Background technique [0002] Prostate cancer is one of the most common cancers in men. Whether a patient has prostate cancer is mainly based on PSA, a cancer factor in the blood. However, since the diagnostic accuracy rate is as low as 30%, a considerable number of patients require additional invasive biopsies, resulting in side effects such as bleeding and pain. [0003] As a non-invasive method, diagnostic testing using urine is convenient for patients and does not require invasive biopsies, thereby diagnosing cancer without side effects. However, the concentration of nucleic acids in urine is low, and there are fewer components of prostate origin, especially the content of prostate-specific RNA is not high. In most cases, R...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6806
CPCC12Q1/6806C12Q2527/125C12Q2527/127
Inventor 徐根明汤链王永利鞠巍冯金儒贺田芬刘雅静
Owner HUNAN YEARTH BIOTECHNOLOGICAL CO LTD
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