Normal-temperature preservative for free DNA in urine

A technology of room temperature preservation and preservative, applied in the field of biomedicine, can solve the problems of inability to stably preserve urine cfDNA, relying on cold chain, etc., and achieve the effects of avoiding self-degradation, inhibiting nuclease, and easy to use

Pending Publication Date: 2020-11-17
合肥铼科生物科技有限公司
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  • Abstract
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  • Application Information

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Problems solved by technology

[0006] In view of this, the present invention provides a urine free DNA preservation agent at room temperature. The preservation solution does not need to rely on the cold chain, and can be stably transported and preserved at room temperature for up to 14 days, effectively preventing the degradation of cfDNA in urine and the release of genomic DNA. At the same time, it solves the problems that current samples rely on cold chain and cannot stably store urine cfDNA, and is suitable for clinical detection needs of targeted cfDNA

Method used

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  • Normal-temperature preservative for free DNA in urine
  • Normal-temperature preservative for free DNA in urine
  • Normal-temperature preservative for free DNA in urine

Examples

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Embodiment 1

[0033] The preparation of embodiment 1 urine free DNA room temperature preservative

[0034] Prepare urine cell-free DNA room temperature preservative according to the following formula:

[0035] preservative components quality score range Recipe 1 Recipe 2 Recipe 3 Recipe 4 Recipe 5 nuclease inhibitor 1-5％ 1 2 2.5 4 5 cell fixative 10-25％ 10 12 15 20 25 cell membrane protector 3-15％ 3 6 10 12 15 Formaldehyde Quencher 1-5％ 1 2 2.5 4 5 PH buffer 50-85％ 85 78 70 60 50

[0036] Wherein, the nuclease inhibitor includes tripotassium edetate, aurintricarboxylic acid and vanadium-ribonucleoside complex, and its mass ratio is 5:3:2; the cell fixative includes diazolidinyl urea, Imidazolidinyl urea, sodium hydroxymethylglycine, its mass ratio is 5:3:2; cell membrane protective agent includes 0.1-0.5M glycine; formaldehyde quencher includes lysine and arginine, its mass ratio is 6 : 4; PH buffer includes 1...

Embodiment 2

[0037] Example 2 The preservation effect verification of urine free DNA normal temperature preservative

[0038] 1. Qubit 4.0 test verifies the preservation effect of urine free DNA room temperature preservative

[0039] 1) Collect 20 mL of mid-morning urine samples from 10 healthy adults. Among them, the experimental group: add 2 mL of urine free DNA room temperature preservative provided by the present invention to 10 mL of urine sample, and the control group: add 2 mL of DEPC water to 10 mL of urine sample.

[0040] 2) Store at room temperature. On the 0th day, the 5th day, the 7th day and the 14th day, 6mL urine samples were taken from each group of samples, and cfDNA was extracted from the urine samples with QIAamp Circulating Nucleic Acid Kit from QIAGEEN. In order to obtain a better extraction effect, the digestion time with proteinase K at 60°C was extended from 30 min to 1 h, and the rest was operated according to the kit instructions, and the elution volume was 20 μ...

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Abstract

The invention provides a normal-temperature preservative for free DNA in urine. The preservative is prepared from a nuclease inhibitor, a cell fixing agent, a cell membrane protective agent, a formaldehyde quenching agent and a PH buffer agent. Through synergistic cooperation of related components, degradation of cfDNA in the urine and release of genomic DNA can be effectively prevented, meanwhile, the problems that a current sample depends on a cold chain and cfDNA in the urine cannot be stably preserved are solved, the preservation effect of stably transporting and preserving the free DNA inthe urine at normal temperature for 14 days is achieved, and the normal-temperature preservative is suitable for the clinical detection requirement for targeted cfDNA.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a urine free DNA preservation agent at room temperature. Background technique [0002] Cancer is the most serious public health problem in the world. Clinically, lesion tissues and cells are mainly used to detect and monitor cancer patients, and the detection and analysis of tissue samples are vulnerable to tumor heterogeneity, sampling difficulties, and high trauma. limit. cfDNA (circulating cell-free DNA) was first reported by Mande et al. in 1948. It was not until the 1990s that the RAS gene fragment was detected from the peripheral blood of tumor patients, and the significance of cfDNA was recognized by people. attach importance to. With the development of liquid biopsy and sequencing technologies, cell-free DNA (cfDNA) or circulating tumor DNA (circulating tumor DNA, ctDNA) are increasingly being used for early screening of cancer, targeted diagnosis and prognostic as...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/10
Inventor 黄服喜刘松李基君陈香
Owner 合肥铼科生物科技有限公司
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