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Novel saliva preserving fluid as well as preparation method and application thereof

A technology for preserving liquid and saliva, applied in the field of molecular biology, can solve the problems of detection inhibition, cumbersome pretreatment, downstream DNA degradation, etc., and achieve the effects of inhibiting growth, convenient preparation, and simplified pretreatment steps

Pending Publication Date: 2020-04-17
申友基因组研究院(南京)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

On the other hand, inappropriate storage solution can cause DNA degradation and inhibition of downstream detection
[0005] Usually the saliva preservation solution on the market requires centrifugation to collect saliva cells and then lysate the cells to lyse the cells, which requires the help of a centrifuge and cumbersome pretreatment

Method used

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  • Novel saliva preserving fluid as well as preparation method and application thereof
  • Novel saliva preserving fluid as well as preparation method and application thereof
  • Novel saliva preserving fluid as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Components and content of saliva preservation fluid: EDTA 10mmol / L, Tris-HCl 20mmol / L, sodium chloride 100mmol / L, sodium diacetate 1% (w / v), proclin300 0.01% (v / v), TWEEN20 1 % (w / v), SDS 1% (w / v), pH value of the system is 8.

[0025] The preparation method is as follows:

[0026] (1) Prepare 0.1mol / L EDTA, Tris-HCl buffer solution with pH 8, and 20% SDS (w / v) storage aqueous solution respectively, and store at room temperature;

[0027] (2) According to the total volume of the pre-prepared saliva for preservation, combined with the content requirements of each component in the system, mix according to the following ratio: 0.1mol / L EDTA aqueous solution 10ml; 0.1mol / L Tris-HCl (PH=8) Buffer 20ml; 20% SDS (w / v) aqueous solution 5ml; TWEEN 20 1ml; proclin300 0.1ml; add 1g sodium diacetate, 0.58g sodium chloride, add sterile water to 100ml, filter out Bacteria, stored at 15-25°C for later use.

[0028] The production and use method of saliva preservation fluid:

[002...

Embodiment 2

[0033] (1) The saliva sample preservation solution of the present invention produced in Example 1 was subjected to a stability test, and 2.5ml of saliva was collected, mixed upside down, and saliva samples from 5 subjects were collected for future use.

[0034] (2) After the saliva samples stored in the saliva preservation solution were placed at room temperature for 0 days, 7 days, 14 days, 21 days, and 30 days, 450ul saliva samples were taken, and Nanjing Shenyou nucleic acid extraction and purification kit (magnetic beads Method)-Type I, extract the genomic DNA of the saliva sample, the total volume of DNA is 80ul. Take 3 ul of saliva DNA samples stored for 0 days and 30 days, add them to 1.0% agarose gel electrophoresis for detection, and use DL15000Marker to mark them. For details, see figure 1 (Samples 1-5 are saliva sample DNA stored for 0 days, and samples 6-10 are saliva sample DNA stored for 30 days); take 2ul saliva DNA and use Qubit to detect its concentration (uni...

Embodiment 3

[0054] The saliva preservation fluid of the present invention produced in Example 1 is detected with the BIOG Elite saliva preservation fluid, and the steps are as follows:

[0055] (1) Take 10 samples of saliva from the subject, shake and mix them evenly and divide them into 2 parts. Preserve 2ml of saliva samples with the above-mentioned saliva preservation composition, and preserve 2ml of saliva samples with 40ul BIOG Elite saliva preservation solution according to the instructions. After the saliva samples were left at room temperature for 14 days, take 450ul saliva samples and use the nucleic acid extraction and purification kit (magnetic bead method) to extract the genomic DNA of the samples. The saliva samples of the present invention are numbered 1-10. Numbered 11-20.

[0056] (2) Detect its concentration with Qubit, see Table 5 specifically, show that the DNA concentration that the saliva preservation liquid of the present invention preserves saliva sample extraction ...

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Abstract

The invention relates to a novel saliva preserving fluid comprising the following components: 1-10mmol / L of EDTA, 1-50mmol / L of Tris-HCl, 10-100 mmol / L of sodium chloride; 0.1-3% (w / v) of sodium sorbate, 0.1-3% (w / v) of sodium diacetate, 0.01-0.1% (v / v) of proclin 300, 0.1-1% (v / v) of TWEEN20, 0.1-1% (w / v) of NP40 and 0.1-2% (w / v) of SDS. The novel saliva preserving fluid is used for preservingsaliva samples, can effectively inhibit nuclease activity, prevent microbial pollution, guarantee DNA fragment integrity and purity, simplify pretreatment steps, is suitable for centrifuge-free and low-temperature environments, can guarantee transport stability of samples among different places, and is suitable for molecular epidemiological large-scale population investigation and research.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and specifically relates to a composition of a novel saliva preservation fluid and methods for its preparation, production and use, which are suitable for transportation and storage after saliva is collected, and are only used for in vitro detection later. Background technique [0002] With the progress of society, nucleic acid DNA is more and more widely used in genetic testing, disease detection and treatment, forensic identification and other fields. Usually, the sample DNA for human genetic analysis comes from blood, tissue, and body fluids. The nucleic acid DNA used for PCR amplification and detection generally comes from blood samples. The collection of blood samples requires professional skills and causes certain trauma to the human body. Blood clots are caused by factors such as blood samples being stored for too long, which can also cause inconvenience in extraction. Non-inva...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/24
CPCC12Q1/24
Inventor 董辉金维荣秦红友杨红
Owner 申友基因组研究院(南京)有限公司
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