CN107267500A - A kind of dissociative DNA preserves liquid and its preparation method and application - Google Patents
A kind of dissociative DNA preserves liquid and its preparation method and application Download PDFInfo
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- CN107267500A CN107267500A CN201710579282.9A CN201710579282A CN107267500A CN 107267500 A CN107267500 A CN 107267500A CN 201710579282 A CN201710579282 A CN 201710579282A CN 107267500 A CN107267500 A CN 107267500A
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Abstract
Liquid, including following components are preserved the invention discloses a kind of dissociative DNA, in terms of 100mL, the content of each component is：8~32g of imidazolidinyl urea；0.005~0.036mol of disodium ethylene diamine tetraacetate；0~2.7g of glycine；The μ L of Triton X 100 50~100；0~0.005mol of pH stabilizers；70~85mL of deionized water.The dissociative DNA, which preserves liquid, intactly medium-term and long-term to preserve human urine, the dissociative DNA in tissue fluid, can be applicable multisample urine, the preservation and detection of tissue fluid dissociative DNA, applied widely, it is ensured that the degree of accuracy of subsequent samples detection and confidence level.
Description
Technical field
The present invention relates to dissociative DNA preservation field, it is specifically related to a kind of dissociative DNA and preserves liquid and preparation method thereof and answer
With.
Background technology
After Apoptosis or necrosis, the DNA of cell interior is released into body fluid formation dissociative DNA (cfDNA).And cancer patient
CfDNA in the cycling tumor DNA (ctDNA) that contains and the generation evolution of tumour be closely related.Dissociative DNA and tumor group
The gene mutation knitted is consistent, and hereditary information is enriched；With tumor load positive correlation, detection sensitivity height；It is earliest to advancing of disease
Make a response；Half-life short (<1.5h), monitoring in real time can be achieved.Therefore, dissociative DNA is used as important tumour-specific mark
Thing, completely acquisition and preservation in terms of cancer early sieve, stage medication guidance and prognosis then to applying offer premise and basis.
In the prior art, dissociative DNA is preserved frequently with blood sample, it is necessary to operated through specialist, process is complicated and consumes
It is time-consuming；Sample component is complicated and preservation extraction difficulty is larger；Dissociative DNA fragment distribution is in 35~1500bp, and tumour
Dissociative DNA fragment focuses mostly in 100~300bp, and fragment is small easy to lose and integrality that lose hereditary information；Free cell is unstable
The intracellular DNA discharged after crushing calmly causes the pollution of dissociative DNA, causes testing result distortion；DNase is abundant in sample causes
Dissociative DNA degraded is quick and is difficult to preserve；Dissociative DNA preserves the cold chain transportation for usually requiring cryogenic conditions, and cost is higher.
The content of the invention
Present invention seek to address that problem as described above.Liquid is preserved it is an object of the invention to provide a kind of dissociative DNA, mainly
Preservation applied to human urine, tissue fluid dissociative DNA.For the mode of traditional evacuation blood collection, urine specimen
Noninvasive, easy to operate, cost is low, be easily accepted by completely for gatherer process, can expand genopathy examination or diagnosis to greatest extent
Sample scope.Tissue fluid generally refers to ascites and hydrothorax, and both samples can be obtained by Minimally Invasive Surgery, and dissociative DNA is compared with blood
Starch that richness is higher, content is more, and can continuously collect, dynamic monitoring tumour with the time Genetic Variations process, and finally
For clinical treatment and diagnosis.The preservation liquid of the present invention, DNA fragmentation preservation integrity degree height, can for a long time be protected in room temperature realization simultaneously
Deposit；Sample Dilution can be at utmost avoided, sample collection, preservation, cost of transportation is finally reduced.
According to an aspect of the present invention, liquid, including following components are preserved the invention provides a kind of dissociative DNA, with
100mL is counted, and the content of each component is：
Wherein, including following components, in terms of 100mL, the content of each component is：
Wherein, including following components, in terms of 100mL, the content of each component is：
Wherein, including following components, in terms of 100mL, the content of each component is：
Wherein, the pH value range that the dissociative DNA preserves liquid is 5.5~7.0.
Wherein, the pH stabilizers can be Tris-HCl buffer solutions, Acetic acid-sodium acetate buffer solution, citric acid-citric acid
Sodium buffer solution.
Wherein, the ratio that the dissociative DNA preserves liquid with preserving sample is 1:20~1:10.
There is provided the preparation method that a kind of dissociative DNA preserves liquid according to another aspect of the present invention, it is characterised in that including
Following steps：The disodium ethylene diamine tetraacetate of predetermined content, pH stabilizers, imidazolidinyl urea, glycine are placed in beaker, 60
~65 DEG C of 25~35min of water-bath；After dissolving, the Triton X-100 of predetermined content are added, deionized water is finally added and is settled to
100mL, is well mixed；0.25 μm of filter membrane suction filtration, produces dissociative DNA and preserves liquid.
Wherein, comprise the following steps：By the disodium ethylene diamine tetraacetate of predetermined content, pH stabilizers, imidazolidinyl urea, sweet
Propylhomoserin is placed in beaker, 65 DEG C of water-bath 30min；After dissolving, the Triton X-100 of predetermined content are added, deionization is finally added
Water is settled to 100mL, is well mixed；0.25 μm of filter membrane suction filtration, produces dissociative DNA and preserves liquid.
Liquid is preserved on preservation urine, tissue fluid dissociative DNA there is provided dissociative DNA according to the third aspect of the present invention
Using.
Dissociative DNA in human urine, tissue fluid sample is extremely unstable, the dirt of cellular content after free cell is broken
Dye, germ contamination, DNase catalytic degradation, pH changes can influence the stabilization of dissociative DNA in sample；Ketoboidies, albumen in sample
Dissociative DNA can also be interfered Deng composition.
The imidazolidinyl urea that the dissociative DNA of the present invention is preserved in liquid can be with stabilizing cell membrane as formaldehyde sustained release agent, it is to avoid
The intracellular DNA of sample is discharged into extracellular, pollution dissociative DNA；Imidazolidinyl urea can efficiently press down as broad-spectrum antibacterial agent simultaneously
Bacterial reproduction processed；It is the main function reagent for stablizing sample dissociative DNA.
The formaldehyde of imidazolidinyl urea sustained release as crosslinking agent/fixative, on the one hand can stablize the cell of sample, albumen,
DNA, but if excessive be possible to cause the reduction of DNA mass.What glycine can be sustained out with imidazolidinyl urea at normal temperatures
Superfluous formaldehyde is combined, and generates hydroxymethyl derivative.Found in research process, for the simple urine of composition, ascites, hydrothorax sample
This, the urine specimen of such as Healthy People, the glycine for adding 0~2.7g can be with imidazolidinyl urea of the constituent content for 8~32g
The superfluous formaldehyde of sustained release is combined so that the dissociative DNA in sample can be preserved well.But for the sample of complicated component,
Such as pregnant woman, sufferer, are had found in experimentation, it is only necessary to which the stable pH of low concentration glycine auxiliary need not be added.
DNase mainly has tri- kinds of DNase I, II, III as the key factor of degraded sample dissociative DNA.Wherein,
DNase I are present in extracellular, and Optimal pH is 7.1, rely on catalytic action of metal ion；DNase II are present in lysosome,
By endocytosis degradation of dna, degradation of dna is independent of metal ion, and optimum pH is 4.5~5.5；DNase III are positioned at
In nucleus, its catalytic dna is degraded independent of metal ion, and optimum pH is 8.5.In the dissociative DNA preservation liquid of the present invention
Adding 0.005~0.036mol disodium ethylene diamine tetraacetate can be pressed down as metal ion chelation agent with complexation of metal ions
DNase I effects processed.
The dissociative DNA of the present invention preserves the content of the pH stabilizers in liquid in 0~0.005mol；Preservation sample can be made
PH is stable 5.5~7.0, can suppress DNase II, the activity of DNase III degradation of dna, Simultaneous Stabilization sample is in normal pH
In the range of, it is ensured that the stabilization of each composition in sample.PH stabilizers can be Tris-HCl buffer solutions, Acetic acid-sodium acetate buffer solution,
The buffer reagents such as citric acid-sodium citrate buffer solution.Meanwhile, glycine can also be common as pH buffer substance and pH stabilizers
Effect, maintains sample pH and each composition stabilization.
The content of disodium ethylene diamine tetraacetate and pH stabilizers in the present invention refers to dry matter content.Preserving liquid
In process for preparation, disodium ethylene diamine tetra-acetic acid solution or Tris-HCl buffer solutions, Acetic acid-sodium acetate buffer solution, lemon are used
Lemon acid-sodium citrate buffer solution.The content of deionized water included in the present invention is gentle for disodium ethylene diamine tetra-acetic acid solution
The summation for the deionized water that the content of the deionized water of fliud flushing and last constant volume are added.
The dissociative DNA of the present invention preserves the Triton X-100 that liquid also added 50~100 μ L content ranges, the content model
The Triton X-100 enclosed can destroy the hydrophilic layer on DNase I surfaces, the activity of DNA enzymatic be reduced, so that in ambient-temp-stable sample
This dissociative DNA.
A series of complex reaction, each group can be produced especially in complex samples in the sample due to preserving each component in liquid
The result of synergy is divided to tend not to precognition.So imidazolidinyl urea, disodium ethylene diamine tetraacetate, pH are stable in the present invention
Agent, Triton X-100, the proportion of glycine are obtained by lot of experiments；And obtained for complicated sample
The preferred proportion scope of each component.So that the preservation liquid of the present invention can be fitted with the dissociative DNA in medium-term and long-term preservation sample
With sample range is wide and preservation effect stable；And the integrity degree of dissociative DNA can be ensured, it is to avoid dissociative DNA is by intracellular DNA
Pollute and interference detection results.
The disodium ethylene diamine tetraacetate of predetermined content, pH stabilizers, imidazolidinyl urea, glycine are placed in beaker, 60
~65 DEG C of 25~35min of water-bath；It is preferred that, 65 DEG C of water-bath 30min；After dissolving, the Triton X-100 of predetermined content are added, most
Addition deionized water is settled to 100mL afterwards, is well mixed；0.25 μm of filter membrane suction filtration, produces dissociative DNA and preserves liquid.
The ratio that this dissociative DNA preserves liquid with preserving sample is 1:20~1:10.In the proportion, the trip in sample
There is good preservation effect from DNA.
This dissociative DNA preserves liquid and is applied to human urine, ascites, the preservation of hydrothorax dissociative DNA, and preservation examination can be made
Agent box, including：Deposit the preservation pipe for thering is the dissociative DNA to preserve liquid, bio-safety bag, specification, personal information card, numbering bar shaped
List is sent in yard, time by special delivery, is assembled into dissociative DNA and is preserved kit；Sample preservation pipe be high-quality it is sterile, without DNase, without RNase and
The 15mL/50mL plastics conical centrifuge tubes of thermal source, include the preservation liquid of about 0.1 times of sample volume；Collection preserves sample and directly turned
Move on to preservation pipe in, closed mixing can room temperature preservation, express delivery list directly arrive pay room temperature transport, customer number bar code facilitate after
Phase sample data is analyzed and processed.
Compared with the prior art, the present invention has the beneficial effect that：
1st, the DNase that the present invention can efficiently suppress in human urine, tissue fluid sample avoids dissociative DNA from being degraded.
2nd, the present invention, which preserves to stablize under liquid room temperature condition, preserves urine, tissue fluid dissociative DNA at least 2 weeks.
3rd, the present invention can stablize the cell in urine specimen, it is ensured that the integrality of cell membrane, it is to avoid dissociative DNA is intracellular
DNA pollution and interference detection results, and the dissociative DNA of the pathology urine specimen such as albuminuria, blood urine also can preserve completely.
4th, preparation method of the present invention is simple, without toxic reagent, without specially treated.
5th, cost is low, be adapted to industrialized production.
The following description for embodiment is read with reference to the drawings, other property features of the invention and advantage will become clear
It is clear.
Brief description of the drawings
The accompanying drawing for being incorporated into specification and constituting a part for specification shows embodiments of the invention, and with
Description is used for the principle for explaining the present invention together, and in the drawings, similar reference is used to represent similar key element, under
Accompanying drawing in the description of face is some embodiments of the present invention, rather than whole embodiments, is come for those of ordinary skill in the art
Say, on the premise of not paying creative work, other accompanying drawings can be obtained according to these accompanying drawings.
Fig. 1 shows that the dissociative DNA of the present invention preserves gel electrophoresis of the liquid to general health human urine sample contrast experiment
Detection figure；
Fig. 2 shows that the dissociative DNA of the present invention preserves gel electrophoresis of the liquid to healthy pregnant women urine specimen contrast experiment and examined
Mapping；
Fig. 3 shows that the dissociative DNA of the present invention preserves liquid and preserves 0d (2h), 8d, 14d detected through gel electrophoresis with control group
Figure；
Fig. 4 shows that the dissociative DNA of the present invention preserves the optical density of band when liquid preserves 0d (2h), 8d, 14d with control group
Distribution map；
Fig. 5 shows that the dissociative DNA of the present invention preserves liquid experimental group, control group urine dissociative DNA sample at 4 DEG C, 37 DEG C
The detected through gel electrophoresis figure of preservation 14 days；
Fig. 6 shows that the dissociative DNA of the present invention preserves liquid experimental group, control group urine dissociative DNA sample at 4 DEG C, 37 DEG C
Preserve the optical density distribution map of 14 days bands；
Fig. 7 shows that the dissociative DNA of the present invention preserves many urine dissociative DNA Sample preservation detected through gel electrophoresis of 14 days of liquid
Figure；
Fig. 8 shows that the dissociative DNA of the present invention preserves the different dilution ratio preservation effect figures of liquid；
Fig. 9 show the dissociative DNA of the present invention preserve liquid and different formulations preserve liquid urine dissociative DNA is preserved 0d ,+
3d ,+8d ,+14d detected through gel electrophoresis figure；
Figure 10 shows that the dissociative DNA of the present invention preserves liquid and different formulations and preserves liquid to 500bp dissociative DNAs anaplasia at any time
The preservation effect of change；
Figure 11 shows that the dissociative DNA of the present invention preserves liquid and different formulations and preserves liquid to 200bp dissociative DNAs anaplasia at any time
The preservation effect of change；
Figure 12 shows that the dissociative DNA of the present invention preserves liquid and different formulations and preserves liquid to 100bp dissociative DNAs anaplasia at any time
The preservation effect of change.
Embodiment
To make the purpose, technical scheme and advantage of the embodiment of the present invention clearer, below in conjunction with the embodiment of the present invention
In accompanying drawing, the technical scheme in the embodiment of the present invention is clearly and completely described, it is clear that described embodiment is
A part of embodiment of the present invention, rather than whole embodiments.Based on the embodiment in the present invention, those of ordinary skill in the art
The every other embodiment obtained under the premise of creative work is not made, belongs to the scope of protection of the invention.Need
Illustrate, in the case where not conflicting, the feature in embodiment and embodiment in the application can be mutually combined.
The present invention provides a kind of dissociative DNA and preserves liquid, including following components, and in terms of 100mL, the content of each component is：Miaow
8~32g of oxazolidinyl urea；0.005~0.036mol of disodium ethylene diamine tetraacetate；0~2.7g of glycine；Triton X-100 50
~100 μ L；0~0.005mol of pH stabilizers；70~85mL of deionized water.
It is preferred that, the content of each component is：15~32g of imidazolidinyl urea；Disodium ethylene diamine tetraacetate 0.02~
0.036mol；0~2.7g of glycine；The μ L of Triton X-100 50~100；0~0.005mol of pH stabilizers；Deionized water 70
~83mL.
It is further preferred that the content of each component is：Imidazolidinyl urea 30g；Disodium ethylene diamine tetraacetate 0.02~
0.036mol；Triton X-100 50μL；PH stabilizers 0.005mol；75~80mL of deionized water.
Or imidazolidinyl urea 30g；Disodium ethylene diamine tetraacetate 0.02mol；Glycine 0.25g；Triton X-100 50μ
L；PH stabilizers 0.005mol；Deionized water 76mL.
Present invention also offers the preparation method that dissociative DNA preserves liquid, comprise the following steps：By the ethylenediamine of predetermined content
Tetraacethyl disodium, pH stabilizers, imidazolidinyl urea, glycine are placed in beaker, 60~65 DEG C of 25~35min of water-bath, it is preferred that
65 DEG C of water-bath 30min；The Triton X-100 of predetermined content are added after dissolving, deionized water is finally added and is settled to 100mL, mix
Close uniform；0.25 μm of filter membrane suction filtration, produces dissociative DNA and preserves liquid.The pH value that gained preserves liquid is 5.5~7.0.
The dissociative DNA of the present invention preserves liquid and is applied to preserve urine, the dissociative DNA of tissue fluid.The preservation liquid and above-mentioned guarantor
The ratio of storage sample product is 1:20~1:10.
Embodiment preserves liquid and prepared
The concentration for measuring predetermined content is the Tris-HCl bufferings that 0.5M disodium ethylene diamine tetra-acetic acid solution, concentration are 1M
Liquid, weighs the imidazolidinyl urea of predetermined content, glycine and is placed in beaker, 65 DEG C of water-bath 30min；After dissolving, addition is predetermined to be contained
The Triton X-100 of amount, finally add deionized water and are settled to 100mL, be well mixed；0.25 μm of filter membrane suction filtration, is produced free
DNA preserves liquid.The data of each component can be as shown in table 1, meanwhile, table 1 also show the pH value that each embodiment preserves liquid.
It is pointed out that the formulation content of the preservation liquid of the present invention is not limited to data in table 1.
The dissociative DNA of table 1 preserves liquid embodiment
Contrast test example
The preservation effect contrast test example that dissociative DNA of the present invention preserves liquid is given below.Due to influence human urine, hydrothorax
And the factor for the DNA stability that dissociates in ascites is basically identical, it is contemplated that obtain the simplicity and diversity of detection sample, selection urine
Liquid sample carries out contrast experiment's test of dissociative DNA preservation.
Urina sanguinis center acid concentration is higher, hereditary information more horn of plenty, and degraded risk is of a relatively high, can more examine preservation liquid matter
Amount, therefore collection urina sanguinis sample.Dissociate DNA content relatively low (content range 0.06~25ng/ μ L, intermediate value in general health human urine
3ng/ μ L), certain difficulty is caused to sample size, extraction operation, dissociative DNA fragment distribution is main in 100~1000bp, therefore
Collecting sample designs a certain amount of DNA marker of addition instantly, facilitates the detection of dissociative DNA preservation effect with presenting, and save the time
With cost.The urine dissociative DNA of following examples preserves in the test experiments of liquid phase pass and gathers urina sanguinis sample simultaneously without specified otherwise
The quantitative exogenous DNA marker of addition.
The general health people of contrast test example 1 and the test of healthy pregnant women urine dissociative DNA contrast experiment preservation effect
1st, instrument, reagent and sample
Key instrument：It is table-type high-speed refrigerated centrifuge (Yancheng City peace letter experimental instruments and equipment limited TGL16M), full-automatic
Instrument for extracting nucleic acid (Hangzhou Ao Sheng Instrument Ltd. Auto Pure-32), the electrophoresis apparatus (bio tech ltd of Beijing 61
DYY-8C), gel imager (Shanghai Tian Neng Science and Technology Ltd.s Tanon 3500), micro-spectrophotometer (Capital Bio
NanoQTM), pipettor etc..
Main agents：Embodiment 1-7 preserve liquid, 0.1 times add certain brand dissociative DNA in blood preserve liquid cw0.1,0.01 times
Add certain brand dissociative DNA in blood and preserve liquid cw0.01, sterilized water control H2O, paramagnetic particle method dissociative DNA extracts kit (English Rui
Sincere biochemical technology (Shanghai) Co., Ltd.).
Sample：General health people, healthy pregnant women freshly voided urine sample；DNA marker(SMOBIO DM2000、
DM2100)。
2nd, experimental program：Quantitative Marker is added into general health people, healthy pregnant women freshly voided urine sample as containing trip
From DNA urine models, embodiment 1-7 is added in proportion and preserves liquid, cw0.1, cw.0.01 preservation liquid, sterilized water, room temperature preservation 1
My god, the dissociative DNA in each group urine specimen is extracted using paramagnetic particle method dissociative DNA extracts kit, electrophoresis detection extracts sample strip
The integrality of band, observes each preservation effect for preserving liquid to urine dissociative DNA.
Deposition condition is：2% Ago-Gel, 1 × TAE buffer solutions, 110V, 25min.Electrophoresis applied sample amount is Marker
0.95 μ L, experimental group DNA extract the μ L of sample 5.Loading order is from left to right：M (DNA Marker), 1 (embodiment 1), 2 (are implemented
Example 2), 3 (embodiments 3), 4 (embodiments 4), 5 (embodiments 5), 6 (embodiments 6), 7 (embodiments 7), 8 (cw0.1), 9
(cw.0.01), 10 (sterilized water).
Experimental group：It is each to preserve liquid (embodiment 1-7 preservations liquid, cw0.1, cw0.01, H2O) room temperature preservation urine dissociative DNA
Model 1 day (+1d)；
Negative control group：Sterilized water H2O group room temperature preservation urines 1 day (+1d).
3rd, experimental result：To general health people, embodiment 1-7, which preserves liquid component, can stablize preservation dissociative DNA+1d；It is right
Embodiment 1,2,6 preserves liquid and the preservation effect that other groups preserve liquid is superior to the preservation effect of dissociative DNA in pregnant woman, experimental group
And sterilized water control.It is specifically shown in Fig. 1,2.
Meanwhile, table 2 also show the UV absorbance detection result to healthy pregnant women urine specimen contrast experiment.
Table 2 respectively preserves the UV absorbance detection result of liquid
| Sample | 260/280 | Concentration (ng/ μ L) | The rate of recovery |
| Embodiment 1 | 1.75 | 12.98 | 90.14% |
| Embodiment 2 | 1.76 | 13.12 | 91.11% |
| Embodiment 3 | 0.00 | -1.12 | 0 |
| Embodiment 4 | 1.70 | 4.62 | 32.08% |
| Embodiment 5 | 1.77 | 7.52 | 52.11% |
| Embodiment 6 | 1.81 | 13.84 | 96.11% |
| Embodiment 7 | 1.77 | 10.39 | 72.15% |
| cw0.1 | 1.66 | 6.30 | 43.75% |
| cw0.01 | 0.00 | -2.46 | 0 |
| H2O | 0.00 | -1.57 | 0 |
Experiment conclusion：Embodiment 1-7 preservation liquid can stablize the dissociative DNA for preserving general health human urine, effect and city
Sell dissociative DNA in blood preservation liquid effect suitable；The preservation liquid phase of embodiment 1,2,6 is compared with other formulas and commercially available dissociative DNA in blood
The dissociative DNA of healthy pregnant women urine can preferably be preserved by preserving liquid, and applicable sample is wider.
Contrast test example 2 preserves liquid and tested with blank control
1st, reagent, instrument and sample
Key instrument：Table-type high-speed refrigerated centrifuge (Yancheng City peace letter experimental instruments and equipment limited TGL16M), magnetic frame
(Aorun Weina New Material Science and Technology Co., Ltd., Shanghai), digital display thermostat water bath (the western instrument science and technology (Shanghai) Co., Ltd. HH- of nation
2), electrophoresis apparatus (the bio tech ltd DYY-8C of Beijing 6 1), gel imager (Shanghai Tian Neng Science and Technology Ltd.s
Tanon 3500), micro-spectrophotometer (Capital Bio NanoQTM), pipettor etc..
Main agents：(English Rui Cheng is biochemical for DNA marker (SMOBIO DM2100), paramagnetic particle method dissociative DNA extracts kit
Scientific and technological (Shanghai) Co., Ltd.), embodiment 6 preserve liquid.
Sample：Gather the fresh urina sanguinis sample about 35mL of general health people.
2nd, experimental program：Urina sanguinis addition preserves room temperature preservation after liquid, 0d (2h), 8d, 14d detection dissociative DNA preservation effect.
Experimental group：14mL urines addition 1mL patents preserve liquid, room temperature preservation after mixing；
Control group：14mL urines add 1mL sterilized waters, room temperature preservation after mixing.
Detection time point：0d(2h)、8d、14d.
Pretreatment：Experimental group, each 700 μ L of two groups of preservation samples of control group are taken, the μ L of equivalent DNA marker 70 are added respectively
In two pipes, fully mix, each group is divided equally in 3 1.5mL centrifuge tube room temperature preservations, for 0d (2h), tri- time points of 8d, 14d
Preservation effect detection.Sample need to carry out 1000 × g centrifugation 10min before detection, take supernatant, use dissociative DNA extracts kit
Carry out dissociative DNA extraction.Electrophoresis detection extracts the integrality of sample strip band, observes each preservation for preserving liquid to urine dissociative DNA
Effect.
Deposition condition is：2% Ago-Gel, 1 × TAE buffer solutions, 110V, 25min.Electrophoresis applied sample amount is Marker 2
μ L, experimental group DNA extract the μ L of sample 7.Loading order (from left to right)：CA-0 (control group 0d), CA-8 (control group 8d), CA-
14 (control group 14d), N-0 (patent preserves liquid group 0d), N-8 (patent preserves liquid group 8d), N-14 (patent preserves liquid group 14d).
3rd, experimental result：Dissociative DNA in liquid product is preserved can completely to be preserved, and the rate of recovery reaches more than 80%, and it is right
Then degraded quickly according to group dissociative DNA, be specifically shown in Fig. 3, Fig. 4.
4th, experiment conclusion：The DNA handled using reagent kit product described in this patent can completely preserve each fragment bar of sample DNA
Band, preservation effect is good.
The extreme room temperature preservation test experiments of the urine specimen of contrast test example 3
1st, instrument, reagent and sample
Key instrument：Full temperature shaken cultivation case (Suzhou Peiying Experimental Equipment Co., Ltd. HZQ-F100), refrigerator (Qing Daohai
Your limited company BCD-190TMPK), table-type high-speed refrigerated centrifuge (Yancheng City peace letter experimental instruments and equipment limited
TGL16M), Full automatic instrument for extracting nucleic acid (Hangzhou Ao Sheng Instrument Ltd. Auto Pure-32), the electrophoresis apparatus (all one's life of Beijing six
Thing Science and Technology Ltd. DYY-8C), gel imager (Shanghai Tian Neng Science and Technology Ltd.s Tanon 3500), micro spectrophotometric
Count (Capital Bio NanoQTM), pipettor etc..
Main agents：Paramagnetic particle method dissociative DNA extracts kit (Ying Ruicheng biochemical technologies (Shanghai) Co., Ltd.), DNA
Marker (SMOBIO DM2000), embodiment 6 preserve liquid.
Sample：General health people's freshly voided urine sample.
2nd, experimental program：General health people's freshly voided urine sample is gathered, 1000 × g centrifugations 10min takes supernatant, and addition is quantitative
DNA Marker are mixed, and experimental group and control group preserve liquid and sterilized water preservation urine, storage temperature condition using patent respectively
For 4 DEG C, 37 DEG C, preserve 0d (2h), 14d and kit extraction and the detection of electrophoretic band integrity degree are carried out to dissociative DNA.
Deposition condition is：2% Ago-Gel, 1 × TAE buffer solutions, 110V, 25min.Electrophoresis DNA extracts sample loading
Measure 7 μ L.
Experimental group (C groups)：Preserve liquid preservation urine specimen and be positioned over 4 DEG C, 37 DEG C；
Control group (N groups)：Sterilized water preserves urine specimen and is positioned over 4 DEG C, 37 DEG C；
3rd, experimental result：Preserve liquid 4 DEG C, 37 DEG C rate of recovery of group more than 80%, and control group then the same day 2h in rapidly
Degraded.It is specifically shown in Fig. 5, Fig. 6.
Table 3 preserves the UV absorbance detection result of liquid and sterilized water
| Sample | 260/280 | Concentration (ng/ μ L) | The rate of recovery |
| C-0d(2h) | 1.75 | 5.01 | 29.47% |
| N-0d(2h) | 1.76 | 16.94 | 99.65% |
| 4℃-C-0d | 0.00 | 0.00 | 0 |
| 4℃-N-14d | 1.70 | 16.68 | 98.12% |
| 37℃-C-0d | 1.77 | 0.07 | 0.41% |
| 37℃-N-14d | 1.81 | 16.78 | 98.71% |
Experiment conclusion：Urine dissociative DNA preserves kit and stable under the conditions of 4 DEG C, 37 DEG C can preserved in urine specimen
Dissociative DNA.
Contrast test example 4 preserves liquid multisample urine dissociative DNA and preserves test experiments
1st, instrument, reagent and sample
Key instrument：It is table-type high-speed refrigerated centrifuge (Yancheng City peace letter experimental instruments and equipment limited TGL16M), full-automatic
Instrument for extracting nucleic acid (Hangzhou Ao Sheng Instrument Ltd. Auto Pure-32), the electrophoresis apparatus (bio tech ltd of Beijing 61
DYY-8C), gel imager (Shanghai Tian Neng Science and Technology Ltd.s Tanon 3500), micro-spectrophotometer (Capital Bio
NanoQTM), pipettor etc..
Main agents：Paramagnetic particle method dissociative DNA extracts kit (Ying Ruicheng biochemical technologies (Shanghai) Co., Ltd.), embodiment
6 preserve liquid；
Sample：General health people's freshly voided urine.
2nd, experimental program：For the presence of individual difference problem, to verify this preservation liquid product to urine dissociative DNA sample
The effective range of protection, gathers random 8 human urine sample, (1# is pregnant women including the people of male 4, the people of women 4；2# is
Menstruating women；3# is marriage and childbirth women；4# is unmarried female；5# is glycosuria male；6#, 8# are that Married Men, 7# are unmarried man
Property), a certain amount of dissociative DNA is added, carries out preserving liquid room temperature preservation 14d, 0d ,+14d dissociative DNAs is extracted.
1000 × g centrifugations 10min obtains sample supernatant, and kit extracts dissociative DNA, and it is free that electrophoresis detection is finally extracted
DNA eluents, deposition condition is：2% Ago-Gel, 1 × TAE buffer solutions, voltage 100V, time 25min.Electrophoresis applied sample amount
The μ L of sample 7 are extracted for experimental group DNA.
3rd, experimental result：The experimental group rate of recovery is more than 80%, and band preserves complete, specific as shown in Figure 7.
4th, experiment conclusion：Crowd's diversity can be met by preserving liquid product, long in being carried out to a variety of urine specimen dissociative DNAs
Phase preserves.
Contrast test example 5 preserves the experiment of liquid fungistatic effect
1st, instrument, reagent and sample
Key instrument：Biohazard Safety Equipment (Dong Lianhaer experimental instruments and equipment limited CLASS II TYPE A2), full Wen Zhen
Swing incubator (Suzhou Peiying Experimental Equipment Co., Ltd. HZQ-F100), pipettor, batch cultur ware, glass spreading rod etc..
Main agents：LB solid mediums, embodiment 6 preserve liquid, sterilized water.
Sample：General health people's freshly voided urine
2nd, experimental program：If experimental group preserves liquid, control group sterilized water and preserves urine specimen, room temperature preservation is taken respectively
6d, 10d urine specimen carry out fungistatic effect detection simultaneously.LB solid mediums spread LB flat boards, take the μ L of each group 200 to preserve sample
Uniformly it is applied on LB flat boards, flat board bacterial plaque number is counted every other day.
Experimental group：Preserve liquid and preserve urine specimen 6d, 10d；
Control group：Sterilized water preserves urine specimen 6d, 10d.
3rd, experimental result：Control group 6d, 10d LB flat boards cover with bacterium colony, experimental group 6d LB flat boards only 6 bacterial plaques,
10d is without bacterial plaque.
4th, experiment conclusion：Preserve liquid fungistatic effect substantially, influence of the bacterial reproduction to dissociative DNA can be avoided.
Contrast test example 6 preserves liquid to be influenceed on albuminuria, conventional 14 Detection results of blood urine mould
1st, instrument, reagent and sample
Key instrument：Urine Analyzer (Yantai Proview Medical Devices Co., Ltd.)；
Main agents：Embodiment 6 preserves liquid, BSA (bovine serum albumin(BSA)/new fresh poultry blood)；
Sample：General health people's freshly voided urine sample.
2nd, experimental program：1g BSA are added into general health people's freshly voided urine sample and are used as albuminuria model；Add 1mL
New fresh poultry blood is as blood urine model, and using liquid is preserved to Sample preservation 8d, 14d (blood urine model is 18d), observation preserves liquid to urine
The impact effect of 14 detection parameters of conventional detection.
Experimental group：Preserve liquid and preserve urine group 0d, 8d, 14d (blood urine 18d)；
Control group：Sterilized water group preserves urine group 0d, 8d, 14d (blood urine 18d)；
3rd, experimental result：It is unchanged that false positive, other data occurs in control group nitrite；Experimental group is preserved after liquid preservation
14 item datas are unchanged.
4th, experiment conclusion：Preservation albuminuria, conventional 14 testing result of blood urine can be stablized by preserving liquid.
Contrast test example 7 preserves experiment of the liquid dilution ratio to urine dissociative DNA preservation effect
1st, instrument, reagent and sample
Key instrument：It is table-type high-speed refrigerated centrifuge (Yancheng City peace letter experimental instruments and equipment limited TGL16M), full-automatic
Instrument for extracting nucleic acid (Hangzhou Ao Sheng Instrument Ltd. Auto Pure-32), the electrophoresis apparatus (bio tech ltd of Beijing 61
DYY-8C), gel imager (Shanghai Tian Neng Science and Technology Ltd.s Tanon 3500), micro-spectrophotometer (Capital Bio
NanoQTM), pipettor etc..
Main agents：Embodiment 6 preserves liquid.
Sample：General health human urine sample；DNA marker(SMOBiO DM2000).
2nd, experimental program：Quantitative Marker is added into general health people's freshly voided urine sample and is used as urine containing dissociative DNA
Model, experimental group presses 1:10、1:20、1:30 ratios addition patent preserves liquid, and room temperature preservation 0d, 7d, 14d observe different dilutions
The patent of ratio preserves preservation effect of the liquid to urine dissociative DNA.
3rd, experimental result：According to 1:10 and 1:The patent of 20 ratios addition preserves that liquid energy is enough efficiently to be preserved in 14 days in room temperature
Urine dissociative DNA, and by 1:Preservation effect of the preservation liquid of 30 ratios addition at 14 days is poor, is specifically shown in Fig. 8.
4th, experiment conclusion：This patent preserves the use dilution ratio of liquid 1:20~1:Between 10.
Contrast test example 8 preserves liquid and other experiments of preservation liquid to urine dissociative DNA preservation effect
1st, instrument, reagent and sample
Key instrument：It is table-type high-speed refrigerated centrifuge (Yancheng City peace letter experimental instruments and equipment limited TGL16M), full-automatic
Instrument for extracting nucleic acid (Ying Ruicheng biochemical technologies (Shanghai) Co., Ltd. EP300), the electrophoresis apparatus (bio tech ltd of Beijing 61
DYY-8C), gel imager (Shanghai Tian Neng Science and Technology Ltd.s Tanon 3500), micro-spectrophotometer (Capital Bio
NanoQTM), pipettor etc..
Main agents：Paramagnetic particle method dissociative DNA extracts kit (Ying Ruicheng biochemical technologies (Shanghai) Co., Ltd.), embodiment
6 preservation liquid, other each compositions preservation liquid, which are shown in, sets group.
Sample：General health human urine sample；DNA marker(SMOBIO DM2000).
2nd, experimental program：Quantitative DNA Marker are added into general health people's freshly voided urine sample as containing dissociative DNA
Urine model, experimental group sets group by liquid composition difference is preserved, and room temperature preservation 0d, 3d, 8d, 14d observe different preservation liquid to urine
The stability of the preservation effect of liquid dissociative DNA and conventional item testing result.
1000 × g centrifugations 10min obtains sample supernatant, and kit extracts dissociative DNA, and it is free that electrophoresis detection is finally extracted
DNA eluents, deposition condition is：2% Ago-Gel, 1 × TAE buffer solutions, voltage 100V, time 25min.Electrophoresis applied sample amount
The μ L of sample 8 are extracted for the μ L of Marker 1.8 (representing 100% rate of recovery), experimental group DNA.
If group：CA (control group-sterilized water), TDZ (certain brand commercially available urine preserve liquid), a, D, E, F (correspondence preserve liquid a,
D, E, embodiment 6 preserve liquid).
CA：Sterilized water, pH8.0；
a:250mM EDTA2Na, 0.9%NaCl, 0.5%PFA, 0.2%DTT, 30% absolute ethyl alcohol 50mM Tris-
HCl, pH8.0；
D：1%PEG 6000,1% butyramide, 250mM EDTA2Na, 50mM Tris-HCl, pH8.0；
E：2%PEG, 0.4%PFA, 20mM EDTA2Na, 5%Tris-HCl, 50% absolute ethyl alcohol, pH7.4；
F：30g imidazolidinyl ureas, 0.02mol EDTA, 50 μ L Triton X-100,0.005mol Tris-HCl, 76mL
Deionized water, pH6.7；
Time point：0d(+4h)、3d、8d、14d.
3rd, experimental result：
a：Each formula preserve the liquid DNA sample detected through gel electrophoresis Fig. 9 and 500bp for representing dissociative DNA typical segments scope,
200bp, 100bp optical density Figure 10-12,500bp represent the larger fragment of dissociative DNA, and 200bp, 100bp represent small fragment.Nothing
Bacterium water CA groups just degraded in 2h 70%, electrophoretic band obscure, it is degradable at 3 days；D groups (3d) can preserve free in short term
DNA, afterwards large fragment be gradually degraded as small fragment (during 14d, C group 500bp contents decline and 200bp, 100bp rise)；A, E exist
Larger fragment (500bp more than 80%) can be completely preserved in experimental period, but below 200bp small fragment can not be preserved；It is commercially available
The preservation liquid F groups of like product TDZ groups and the present invention can be preserved in 14 days to all fragments, but to small fragment 100bp
Preservation effect F groups (about 85%) be substantially better than TDZ groups (about 70%).
b：At the 3rd day of the stability experiment of conventional 14 testing result, there is the false sun of occult blood in sterilized water CA groups
Property, pH increase to 7.5 from 6.0；It is 7.5 that nitrite false positive, pH, which occur, in commercial like product TDZ groups；Patent preserves liquid F groups
The stable only pH of every result increases to 6.5 from 6.0 and slightly changed.Conventional item test experience result only embodies 0 ,+3d ,+8d ,+
14d experimental results are consistent with+3d.
4th, experiment conclusion：Patent preserves liquid phase can more completely, for longer periods compared with certain commercially available brand urine preservation liquid
Preserve ucfDNA and stablize the testing result of routine urinalysis.
In summary, the dissociative DNA provided according to the present invention, which preserves liquid, intactly medium-term and long-term to preserve human urine, group
Knit the dissociative DNA in liquid, multisample urine, the preservation of tissue fluid dissociative DNA can be applicable, it is applied widely, it is ensured that after
The degree of accuracy of continuous pattern detection and confidence level.
Finally it should be noted that：Herein, term " comprising ", "comprising" or its any other variant are intended to non-
It is exclusive to include, so that process, method, article or equipment comprising a series of key elements not only include those key elements,
But also other key elements including being not expressly set out, or also include solid by this process, method, article or equipment
Some key elements.In the absence of more restrictions, the key element limited by sentence " including one ... ", it is not excluded that including institute
Also there is other identical element in process, method, article or the equipment of stating key element.
The above embodiments are merely illustrative of the technical solutions of the present invention, rather than its limitations.Although with reference to the foregoing embodiments
The present invention is described in detail, it will be understood by those within the art that：It still can be to foregoing each implementation
Technical scheme described in example is modified, or carries out equivalent substitution to which part technical characteristic；And these modification or
Replace, the essence of appropriate technical solution is departed from the spirit and scope of various embodiments of the present invention technical scheme.
Claims (10)
1. a kind of dissociative DNA preserves liquid, it is characterised in that including following components, in terms of 100mL, the content of each component is：
2. dissociative DNA as claimed in claim 1 preserves liquid, it is characterised in that including following components, in terms of 100mL, described each
The content of component is：
3. dissociative DNA as claimed in claim 2 preserves liquid, it is characterised in that including following components, in terms of 100mL, described each
The content of component is：
4. dissociative DNA as claimed in claim 2 preserves liquid, it is characterised in that including following components, in terms of 100mL, described each
The content of component is：
5. the dissociative DNA as described in claim any one of 1-4 preserves liquid, it is characterised in that the dissociative DNA preserves the pH of liquid
It is 5.5~7.0 to be worth scope.
6. dissociative DNA as described in claim any one of 1-5 preserves liquid, it is characterised in that the pH stabilizers can be
Tris-HCl buffer solutions, Acetic acid-sodium acetate buffer solution, citric acid-sodium citrate buffer solution.
7. the dissociative DNA as described in claim any one of 1-6 preserves liquid, it is characterised in that the dissociative DNA preserves liquid with protecting
The ratio of storage sample product is 1:20~1:10.
8. a kind of dissociative DNA as described in claim any one of 1-7 preserves the preparation method of liquid, it is characterised in that including with
Lower step：The disodium ethylene diamine tetraacetate of predetermined content, pH stabilizers, imidazolidinyl urea, glycine are placed in beaker, 60~
65 DEG C of 25~35min of water-bath；After dissolving, the Triton X-100 of predetermined content are added, deionized water is finally added and is settled to
100mL, is well mixed；0.25 μm of filter membrane suction filtration, produces dissociative DNA and preserves liquid.
9. dissociative DNA as claimed in claim 8 preserves the preparation method of liquid, it is characterised in that comprise the following steps：Will be predetermined
Disodium ethylene diamine tetraacetate, pH stabilizers, imidazolidinyl urea, the glycine of content are placed in beaker, 65 DEG C of water-bath 30min；It is molten
Xie Hou, adds the Triton X-100 of predetermined content, finally adds deionized water and is settled to 100mL, is well mixed；0.25 μm of filter
Film suction filtration, produces dissociative DNA and preserves liquid.
10. a kind of dissociative DNA as described in claim any one of 1-7 preserves liquid on preservation urine, tissue fluid dissociative DNA
Using.
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Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107881213A (en) * | 2017-11-10 | 2018-04-06 | 人和未来生物科技（长沙）有限公司 | A kind of urine preserves reagent and its application |
| CN107980763A (en) * | 2017-11-10 | 2018-05-04 | 人和未来生物科技（长沙）有限公司 | A kind of excrement preserves reagent and its application |
| CN108048529A (en) * | 2017-12-28 | 2018-05-18 | 广州市金圻睿生物科技有限责任公司 | A kind of quality-control product of lung cancer methylated genes detection kit for stablizing preservation and application |
| CN108823286A (en) * | 2018-07-18 | 2018-11-16 | 北京迈基诺基因科技股份有限公司 | A kind of protective agent and vacuum blood collection tube of dissociative DNA in blood |
| CN109757466A (en) * | 2018-10-29 | 2019-05-17 | 中国医学科学院阜外医院 | Urine saves liquid, urine capture container, method and kit |
| CN110250159A (en) * | 2019-05-28 | 2019-09-20 | 天津中科生物科技有限公司 | A kind of Cell protective solutions and its preparation method and application |
| CN111902545A (en) * | 2019-01-03 | 2020-11-06 | 杭州诺辉健康科技有限公司 | Composition and method for urine sample preservation and DNA extraction |
| CN112662662A (en) * | 2021-01-05 | 2021-04-16 | 南昌艾迪康医学检验实验室有限公司 | Free DNA preservation reagent and preparation method thereof |
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| WO2021195768A1 (en) * | 2020-03-30 | 2021-10-07 | Dna Genotek Inc. | Stabilizing composition and method for preserving a bodily fluid |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013123030A3 (en) * | 2012-02-13 | 2013-10-31 | Streck, Inc. | Blood collection device for stabilizing cell -free plasma dna |
| CN103789202A (en) * | 2014-01-26 | 2014-05-14 | 付士明 | Container for collecting nucleic acid preserved at normal temperature |
| US20160257995A1 (en) * | 2015-03-05 | 2016-09-08 | Streck, Inc. | Stabilization of nucleic acids in urine |
| CN105985904A (en) * | 2015-02-26 | 2016-10-05 | 付士明 | Circulating free DNA vacuum collection tube |
| CN106244535A (en) * | 2016-08-30 | 2016-12-21 | 成都瑞琦科技实业股份有限公司 | The preservative agent of fetal cell-free DNA in maternal plasma and the vacuum test tube of composition thereof |
-
2017
- 2017-07-17 CN CN201710579282.9A patent/CN107267500A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2013123030A3 (en) * | 2012-02-13 | 2013-10-31 | Streck, Inc. | Blood collection device for stabilizing cell -free plasma dna |
| CN103789202A (en) * | 2014-01-26 | 2014-05-14 | 付士明 | Container for collecting nucleic acid preserved at normal temperature |
| CN105985904A (en) * | 2015-02-26 | 2016-10-05 | 付士明 | Circulating free DNA vacuum collection tube |
| US20160257995A1 (en) * | 2015-03-05 | 2016-09-08 | Streck, Inc. | Stabilization of nucleic acids in urine |
| CN106244535A (en) * | 2016-08-30 | 2016-12-21 | 成都瑞琦科技实业股份有限公司 | The preservative agent of fetal cell-free DNA in maternal plasma and the vacuum test tube of composition thereof |
Non-Patent Citations (1)
| Title |
|---|
| 温贤芳: "《中国核农学》", 31 August 1998, 河南科学技术出版社 * |
Cited By (12)
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| CN107980763A (en) * | 2017-11-10 | 2018-05-04 | 人和未来生物科技（长沙）有限公司 | A kind of excrement preserves reagent and its application |
| CN108048529A (en) * | 2017-12-28 | 2018-05-18 | 广州市金圻睿生物科技有限责任公司 | A kind of quality-control product of lung cancer methylated genes detection kit for stablizing preservation and application |
| CN108823286A (en) * | 2018-07-18 | 2018-11-16 | 北京迈基诺基因科技股份有限公司 | A kind of protective agent and vacuum blood collection tube of dissociative DNA in blood |
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| EP3906319A4 (en) * | 2019-01-03 | 2022-11-16 | Hangzhou New Horizon Health Technology Co., Ltd. | Compositions and methods for urine sample storage and dna extraction |
| CN110250159A (en) * | 2019-05-28 | 2019-09-20 | 天津中科生物科技有限公司 | A kind of Cell protective solutions and its preparation method and application |
| WO2021195768A1 (en) * | 2020-03-30 | 2021-10-07 | Dna Genotek Inc. | Stabilizing composition and method for preserving a bodily fluid |
| CN112662662A (en) * | 2021-01-05 | 2021-04-16 | 南昌艾迪康医学检验实验室有限公司 | Free DNA preservation reagent and preparation method thereof |
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