CN109295201A - Predict the method and DNA methylation marker of weightless bone loss bone alkaline phosphatase BALP downside risk - Google Patents
Predict the method and DNA methylation marker of weightless bone loss bone alkaline phosphatase BALP downside risk Download PDFInfo
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- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
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- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
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- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/154—Methylation markers
Abstract
The present invention provides the DNA methylation marker for predicting weightless bone loss bone alkaline phosphatase BALP downside risk, including following one or more genes, site annotation information are shown in Table 3.The present invention also provides corresponding detection methods.The present invention passes through to haemocyte DNA methylation level and serum skeleton metabolism (bone alkaline phosphatase, BALP excavation), it was found that horizontal by detection haemocyte DNA methylation, it can be realized the prediction to human body bone alkaline phosphatase decline individual difference after weightlessness, to which screening is more resistant to the individual of below-G conditions, or for carrying out scientific research to bone alkaline phosphatase situation of change.It is higher with the correlation of human body bone metabolism the present invention provides the DNA methylation site for having forecast function to human body bone alkaline phosphatase downside risk after weightlessness set.
Description
Technical field
The invention belongs to physiology technical fields, and the physiological change after being related to weightlessness condition for a period of time specifically relates to
And predict the method and DNA methylation marker of weightless bone loss bone alkaline phosphatase BALP downside risk.
Background technique
Bone alkaline phosphatase (BALP) is the bone specificity isomers of alkaline phosphatase, is found in osteoblast surfaces
A kind of glycoprotein reflects the biosynthesis activity and function status of osteoblast.Bone alkaline phosphatase has proved to be a kind of
Sensitive reliable skeleton metabolism is the specificity ginseng for being mainly used for Rickets early diagnosis and subclinical identification in recent years
Examine index, and the optimal parameter currently used for evaluating human body bone mineralising obstacle.BALP derive from osteoblast, eliminate liver,
The influence of the illness such as kidney, enteron aisle can more accurately reflect bone metabolism situation.When bon e formation is greater than bone resorption, BALP in serum
It is active then obviously increase, therefore it is reflection one of osteoblast activity and the sensitive indicator of bon e formation.Clinically, it is cured in fracture
During conjunction, bon e formation is active, and BALP activity increases；And when causing bone resorption increased disease such as osteoporosis, myeloma,
BALP activity can then reduce.
Weightlessness bone loss is a kind of long-term, lasting progressive variation, and the result of development will lead to osteoporosis, soft group
The generation of the pathological phenomenons such as calcification, kidney stone is knitted, or even gravity occurs and adapts to obstacle again, so that the body for seriously affecting spacefarer is strong
Health and working efficiency.Existing means are difficult to prevent and delay space bone loss.In micro-gravity conditions, the rate of bone loss is super
Cross monthly lose 10 times of bone density of postmenopausal women on ground.The measuring study of bone density shows to stay in 1-6 months spaces
For spacefarer's bone loss amount between 0.9%-1.8%, this ratio is different because of test position difference during staying, and generally recognizes at present
It is 1%-2% for the amount lost of sclerotin and rock salt monthly.Weightlessness bone loss is that bone resorption increases and bon e formation reduces
The result of comprehensive function.Weightlessness bone loss is under the various stress conditions brought by weightless and space travel,
The result to interact between osteocyte, osteoblast and osteoclast.The reduction of function of osteoblast bone caused by weightlessness
It is played a key role in loss.The decline of BALP level means that weightless or simulated weightlessness lower body skeletonization is thin in serum
The decline of cytoactive and bone formation ability.
To sum up, under weightlessness/simulated weightlessness conditions, human body bone alkaline phosphatase (BALP) is horizontal to be reduced and Human osteoblast's cell
Activity reduces and bone density decline is in close relations.In consideration of it, after being subjected to same weightlessness/simulated weightlessness conditions and influencing, human body
The difference of BALP decline level can also reflect that human body leads to the individual difference of bone loss tolerance degree to below-G conditions.Towards manned
Space mission, as can the BALP downside risk before execution task to occupant or subject under simulated weightlessness conditions carries out in advance
It surveys, optimization member is selected, early stage health protection intervention is provided for occupant and human body bone alkaline phosphatase (BALP) is declined
Risk is studied, and all has significant application value.
Summary of the invention
In order to solve the problems in the existing technology, Head down tilt bed rest human experimentation model, sieve are spent in use -6 of the present invention
Select and identify can predict weightlessness after human serum bone alkaline phosphatase downside risk epigenetics index set.
The present invention provides the DNA methylation marker for predicting weightless bone loss bone alkaline phosphatase BALP downside risk, packet
Following one or more genes are included, gene loci annotation information is shown in Table 3.
These are identified by gene symbol (the 11st column) and at least one chromosomal foci (the 8th, 9,10 column) in table
The preferred methylated nucleotide position of gene.These genes are further identified by probe id (column 1), special in these genes
It is not the identification site CpG in their controlling element (in chromosomal foci).
Hereditary bibliography herein is typically referenced to genome human hg38.In table, probe id refers in array
The sequence indicated on platform, each for inquiring the specific site CpG, chromosome (chr), start site (start) and
End locus (end) can uniquely identify first position nt of each probe.
The present invention provides the method for predicting weightless bone loss bone alkaline phosphatase BALP downside risk, and the method is not with disease
For the purpose of the diagnosing and treating of disease, include the following steps: to determine bone alkaline phosphatase risk genes site in Samples subjects
DNA methylation is horizontal, and the methylation level of the gene loci is compared with control sample, to predict that subject is losing
Weight condition bone alkaline phosphatase BALP downside risk；
Wherein one or more genes of the bone alkaline phosphatase risk genes in claim 1；
The control sample is selected from the sample that bone alkaline phosphatase BALP does not decline under below-G conditions；
The method of the determining DNA methylation level can be determining by any method known in the art, the method
Methylating including full-length genome, screening is analyzed, the methylation profiles of chip platform are analyzed, methylation status of PTEN promoter analysis, flight
The analysis of Mass Spectrometer Method (Sequenome), sulphite treated gene order-checking, joint bisulfites restriction enzyme
Analysis, methylation-specific endonuclease digestion and quantitative polyase chain reaction Conjoint Analysis, methylation sensitive high-resolution are molten
Solution curve analysis or pyrosequencing determination method.
The DNA methylation assay reagent that the present invention provides gene described in claim 1 predicts weightless bone loss bone alkali in preparation
Application in acid phosphatase BALP downside risk diagnostic products, the diagnostic products include a kind of or more in detection claim 1
The DNA methylation assay reagent of kind gene, the methyl that the reagent passes through gene described in claim 1 in detection Samples subjects
Change level, to judge whether subject's bone alkaline phosphatase under below-G conditions has downside risk；
Preferably, the reagent is to methylate to screen the methylation figure of analysis, chip platform based on full-length genome
Spectrum analysis, methylation status of PTEN promoter analysis, flight mass spectrum detection (sequenome), sulphite treated gene order-checking
Analysis, joint bisulfites restriction endonuclease analysis, methylation-specific endonuclease digestion and quantitative polyase chain reaction
Conjoint Analysis, methylation sensitive high-resolution melting curve analysis or pyrosequencing test and analyze required reagent.
As another preferred embodiment, the methylation level of the gene is in the upstream of the open reading frame of marker gene
What region, especially promoter region determined；Or one of following:
(a) nucleic acid defined in the chromosomal foci as shown in claim 1；
(b) comprising the site CpG of nucleic acid a；
(c) nucleic acid (a) being no more than with above-mentioned nucleic acid a distance in 1000 nucleotide.
As another preferred embodiment, the reagent including specific amplification include methylation sites including claim 1
Described in gene primer；
As further optimisation, the diagnostic products are kit, chip or microarray dataset.
The present invention provides one group of nucleic acid primer and/or hybridization probe, to one or more gene described in claim 1
Potential methylation region be specific；
Preferably, the primer and/or hybridization probe are specific for amplification marker gene open reading frame
Methylate upstream region, especially promoter region；Or methylation is one of following:
(a) nucleic acid defined in the chromosomal foci as shown in claim 1；
(b) comprising the site CpG of nucleic acid a；
(c) nucleic acid (a) being no more than with above-mentioned nucleic acid a distance in 1000 nucleotide.
Primer and/or probe are used to target a potential first in the DNA molecular of the one or more genes selected in table 3
Base region.
The present invention provides a kind of kit, and the kit includes one group of nucleic acid primer as claimed in claim 6 or hybridization
Probe further includes methylation-specific restriction enzyme and/or takes off the reagent of amine for bisulfites nucleotide.
The present invention provides a kind of diagnostic products for predicting weightless bone loss bone alkaline phosphatase BALP downside risk, described to examine
Stopping pregnancy product include the reagent for being able to detect the level of gene methylation described in claim 1；
Preferably, the reagent of gene methylation level described in the detection claim 1 includes being used for full base
Because group methylation screening analysis a reagent, for based on chip methylation profiles analysis reagent, be used for methylation-specific
The reagent of PCR analysis is surveyed for the reagent of flight mass spectrum detection (sequenome), for sulphite treated genome
The reagent of sequence analysis, is used for methylation-specific restriction endonuclease at the reagent for combining bisulfites restriction endonuclease analysis
The reagent of digestion and quantitative polyase chain reaction Conjoint Analysis, for the examination of methylation sensitive high-resolution melting curve analysis
Agent or the reagent tested and analyzed for pyrosequencing.
As another preferred embodiment, the reagent including specific amplification include methylation sites including claim 1
Described in genetic fragment primer；
As another preferred embodiment, the diagnostic products are kit, chip or microarray dataset.
The present invention provides the method for prediction weightlessness bone loss bone alkaline phosphatase BALP decline, and the method is not with disease
For the purpose of diagnosing and treating, steps are as follows:
(1) DNA methylation for measuring DNA methylation site is horizontal；
(2) building of two disaggregated model of logistic regression is carried out, and substitutes into following formula:
Wherein, X=intercept coefficient+DNA methylation site DNA methylation level × independent variable coefficient, Y are bone alkalinity phosphorus
Sour enzyme BALP downside risk score；Such as Y < 0.5, then it is determined as that bone alkaline phosphatase BALP declines.
The present invention passes through to haemocyte DNA methylation level and serum bone density index (bone alkaline phosphatase, BALP)
It excavates, discovery is horizontal by detection haemocyte DNA methylation, can be realized to human body bone alkaline phosphatase decline individual after weightlessness
The prediction of difference, thus individual of the screening more resistant to below-G conditions, or for being carried out to bone alkaline phosphatase situation of change
Scientific research.
The present invention provides the DNA methylation sites for having forecast function to human body bone alkaline phosphatase downside risk after weightlessness
Set is higher with the correlation of human body bone metabolism.This is found to be system and discloses human body bone loss caused by below-G conditions influence
New clue is provided, further investigation is worth.
Specific embodiment
Embodiment below facilitates a better understanding of the present invention, but does not limit the present invention.Experiment in following embodiments
Method is unless otherwise specified conventional method.Test material as used in the following examples is unless otherwise specified city
It sells.
The present invention provides the DNA methylation marker column that the human body bone alkaline phosphatase downside risk after weightlessness can be predicted
Table (gene location annotation information and estimated performance are shown in Table 3), and provide a kind of prediction it is weightless after under human body bone alkaline phosphatase
The method of risk drops.In a specific embodiment, pass through the methylation level for the marker cg22433798 that methylates, evaluation
Human body bone alkaline phosphatase declines individual difference, the subject of identification bone alkaline phosphatase decline after weightlessness, and then screens more
It is resistant to the individual of below-G conditions.
Although the present invention, which is regardless of, is limited to any theory, inventor thinks methylation marker levels and weightlessness in table 3
Under the conditions of there are correlativities between bone alkaline phosphatase downside risk.
In a specific embodiment in the present invention, bone alkaline phosphatase downside risk score Y, DNA methylation water
The relationship of flat cg22433798 meets
Wherein, X=5161+cg22433798 × (- 5421), the cg22433798 in formula are the DNA in specific gene site
Methylation level.Such as Y < 0.5, then it is determined as that bone alkaline phosphatase declines.
Embodiment 1
1, -6 degree Head down tilt bed rest simulated weightlessness model
(1) volunteer
Testing volunteer is 15 healthy non-smoking males, special secondary school and the above schooling of special secondary school.Age was at 22 years old to 34
Between year (M=26.62, SD=4.21), height (M=170.31, SD=4.02) between 160cm to 174cm, weight 53~
72Kg (M=62.81, SD=5.90).It without special medical history, eyesight or corrects defects of vision normal, is dextro manuality, non athlete.Institute
Some test volunteers fill in informed consent form before receiving experiment.
(2) experiment condition
15 volunteers downward -6 degree state of holding head, continuous life 45 days on -6 degree bed for lying on.It is held in experimentation
Continuation of insurance holds body axial -6 and spends the downward state in head.
(3) experimental arrangement
Bed rest experiment starts first 1 day, acquires 15 subject's blood cell samples by EDTA anticoagulant tube, freezes in -80 DEG C
Refrigerator remains subsequent DNA methylation chip detection；Subject 1 day and bed the before bed test are detected by elisa technique
45 days serum bone alkaline phosphatase indexs.
2, data acquire overall situation
It is as shown in table 1 that data acquire situation.
1 data of table acquire situation table
3, bone alkaline phosphatase BALP detection method and quality control
Using BALP in ELISA method detection serum, using people's alkaline phosphatase in double antibody sandwich method measurement sample
(ALP) horizontal.Use Human BALP (Bone Alkaline Phosphatase) ELISA reagent of Immunoway company
Box.Be coated with microwell plate with people's alkaline phosphatase (ALP) antibody of purifying, solid phase antibody be made, toward in the micropore for being coated with monoclonal antibody according to
Secondary addition alkaline phosphatase (ALP), then in conjunction with alkaline phosphatase (ALP) antibody of HRP label, form antibody-antigene-enzyme mark
Antibody complex, after thoroughly washing plus substrate TMB develops the color.TMB converts au bleu under the catalysis of HRP enzyme, and in acid
Final yellow is converted under effect.Alkaline phosphatase (ALP) in the depth and sample of color is positively correlated.Existed with microplate reader
Absorbance (OD value) is measured under 450nm wavelength, people's alkaline phosphatase (ALP) concentration in sample is calculated by standard curve.
After degree Head down tilt bed rest test in 45 days -6,6 subject's bone alkaline phosphatases decline, and relative drop amplitude is
4.4%-49.9%；9 subjects rise.Because the decline of bone alkaline phosphatase level in serum means that body osteoblast is living
Property and bone formation ability decline, therefore by subject be divided into bone alkaline phosphatase decline high risk group (H group, relative drop width
Degree amounts to 6 subjects more than 4%) and low-risk group (L group, bone alkaline phosphatase level is non-decreasing, amounts to 9 subjects).
The variation tendency of 2 bone alkaline phosphatase BALP of table
4, DNA methylation express spectra detects
It is detected using Illumina Human 450k DNA methylation chip.
In view of biochip technology relative maturity, and there is the commercial technologies service platform of operation maturation in the country, using outer
The mode of association's test completes Illumina Human 450k DNA methylation chip testing.Main technologies include:
(1) genome DNA is extracted.It is extracted using commercial kit QIAamp DNA Mini Kit kit total
DNA。
(2) DNA quality inspection.Using NanoDrop and Agrose Gel quality detecting method, pass through A260/280 signal, A260/230
The quality inspections parameters such as signal, DNA total amount (μ g) extract the quality control of DNA.
(3) bisulfite conversion is handled using Zymo EZ DNA Methylation Kit kit.
(4) WGA mode expand, fragmentation, using Illumina chip scanner, with GenomeStudio software control.
(5) chip hybridization, elution, extension, imaging, using GenomeStudio software.
(6) data are analyzed.
Above-mentioned experimental implementation is carried out by the standard practice instructions of related kit and company.
5, DNA methylation chip of expression spectrum data prediction
Using the Illumina Human 450k DNA methylation Chip data output file * .idat of standard
In file, data analysis is carried out using R language platform (v3.4.4) and Rstudio tool (v1.1.442).Pre-process institute's recruitment
Tool includes: minfi (v1.24.0), ChAMP (v2.9.10).By filtering following probe: (1) detecting p value and be greater than 0.01；
(2) be more than 5% sample middle probe bead-count number less than 3；(3) probe of non-CG beginning；(4) site SNPs starts
Probe；(5) probe in multiple gene locations is annotated；(6) probe of the annotation on X and Y chromosome, finally obtains 412345
Probe, 15 subjects DNA methylation express modal data, be used for subsequent analysis.
6, Head down tilt bed rest simulated weightlessness conditions lower lumbar spine bone loss risk profile methylation sites screen
(1) aspect of model screening 1 --- standard deviation filtering
Standard deviation 15 subjects is calculated between each probe, probe is ranked up by standard deviation, is filtered out
The probe that benchmark methylation level differs greatly in subject population.Screening threshold value is set as sd > 0.02, thus filters out
117872 probes are used for subsequent analysis.
(2) aspect of model screening 2 --- differential expression screening
To 117872 methylation probes, between bone alkaline phosphatase decline high risk group (H group) and low-risk group (L group)
Carry out t check analysis.It is threshold value with p-value < 0.05, the probe of methylation level significant difference between screening group.Thus it screens
2487 probes out are used for subsequent analysis.
(3) building of unit point prediction model and performance evaluation of logic-based regression model
Using Logic Regression Models, disaggregated model building is carried out respectively with each site in above-mentioned 2487 probes.Using staying
One verification method evaluates the performance of each site model, output model AUC, Odds-Ratio value, model accuracy Accuracy, model
The parameter informations such as precision confidence interval and p value.It is threshold value with p-value < 0.05, filtering out 133 has higher forecasting performance
Methylation sites.Wherein 90 site annotations are on known (table 3).
Table 3 there is the methylation characteristic site of predictive ability to gather bone alkaline phosphatase downside risk under below-G conditions
Function enrichment analysis is done for gene where above-mentioned 90 probes filtered out using Logic Regression Models.In table 4
Analysis is the results show that gene where having the methylation sites of predictive ability to bone alkaline phosphatase downside risk under below-G conditions
Most significant enrichment is in Chemokine signaling pathway signal path (p-value=1.22e-2).
The function of gene is enriched with result where 4 below-G conditions lower lumbar spine bone loss risk profile methylation sites of table
(4) bone alkaline phosphatase BALP downside risk prediction model example before and after simulated weightlessness
It is input with methylation sites cg22433798 in table 3.Probe cg22433798 homologue sequence and corresponding
Location information is following (italic overstriking font shows that sequence includes probe location upstream and downstream 25bp nucleic acid sequence), with reference to genome
For human hg38:
> chr7:584994-585045 (cg22433798)
Bone alkaline phosphatase is declined into high risk group (H group, relative drop amplitude are more than 4%, amount to 6 subjects) label
Classifier is " 0 ", and low-risk group (L group, bone alkaline phosphatase level is non-decreasing, amounts to 9 subjects) labeled bracketing symbol is
"1".Two disaggregated model of logistic regression is carried out using R language platform (v3.4.4) and kit caret (v6.0.80) to construct, it is right
Bone alkaline phosphatase decline high risk group and low-risk group are predicted under below-G conditions.Prediction model coefficient such as table 5.
5 prediction model coefficient of table
| Variable | Coefficient |
| Cg22433798 (independent variable) | -5421 |
| Chang Bianliang | 5161 |
Calculation formula used is as follows:
Wherein, X=5161+cg22433798 × (- 5421), cg22433798 are the DNA methylation in specific gene site
Level, Y are bone alkaline phosphatase downside risk score.Such as Y < 0.5, then it is determined as that bone alkaline phosphatase declines.
By staying a verifying to analyze, prediction model AUC=1 (p-value=4.7e-04), other model performance parameters
See Table 6 for details for output.The result shows that the prediction model can concentrate bone density high risk/low-risk crowd progress accurate pre- to data
It surveys.
6 prediction model performance parameter of table
Finally, it should be noted that the foregoing is only a preferred embodiment of the present invention, it is not intended to restrict the invention,
Although the present invention is described in detail referring to the foregoing embodiments, for those skilled in the art, still may be used
To modify the technical solutions described in the foregoing embodiments or equivalent replacement of some of the technical features.
All within the spirits and principles of the present invention, any modification, equivalent replacement, improvement and so on should be included in of the invention
Within protection scope.
Claims (10)
1. predict the DNA methylation marker of weightless bone loss bone alkaline phosphatase BALP downside risk, including following one or
Multiple genes, site annotation information are as follows:
2. the method for predicting weightless bone loss bone alkaline phosphatase BALP downside risk, the method with the diagnosis of disease and are not controlled
For the purpose for the treatment of, it is characterised in that: include the following steps: to determine bone alkaline phosphatase risk genes site in Samples subjects
DNA methylation is horizontal, and the methylation level of the gene loci is compared with control sample, to predict that subject is losing
Weight condition bone alkaline phosphatase BALP downside risk；
Wherein one or more genes of the bone alkaline phosphatase risk genes in claim 1；
The control sample is selected from the sample that bone alkaline phosphatase BALP does not decline under below-G conditions；
Preferably, the method for the determining DNA methylation level includes full-length genome methylation screening analysis, chip platform
Methylation profiles analysis, methylation status of PTEN promoter analysis, flight mass spectrum detection (Sequenome), sulphite treated base
Because of a group sequencing analysis, joint bisulfites restriction endonuclease analysis, methylation-specific endonuclease digestion and quantitative polymerization
Enzyme chain reaction Conjoint Analysis, methylation sensitive high-resolution melting curve analysis or pyrosequencing determination method.
3. the DNA methylation assay reagent of gene described in claim 1 predicts weightless bone loss bone alkaline phosphatase BALP in preparation
Application in downside risk diagnostic products, it is characterised in that: the diagnostic products include one or more in detection claim 1
The DNA methylation assay reagent of gene, the methylation that the reagent passes through gene described in claim 1 in detection Samples subjects
Level, to judge whether subject's bone alkaline phosphatase under below-G conditions has downside risk；
Preferably, the reagent is methylated based on full-length genome screening analysis, the methylation profiles analysis of chip platform, first
The analysis of base specific PCR, flight mass spectrum detection (sequenome), the analysis of sulphite treated gene order-checking, joint
Bisulfites restriction endonuclease analysis, methylation-specific endonuclease digestion and quantitative polyase chain reaction Conjoint Analysis,
Methylation sensitive high-resolution melting curve analysis or pyrosequencing test and analyze required reagent.
4. application according to claim 3, it is characterised in that: the methylation level of the gene is opening in marker gene
Put what the upstream region of reading frame, especially promoter region determined；Or one of following:
(a) nucleic acid defined in the chromosomal foci as shown in claim 1；
(b) comprising the site CpG of nucleic acid a；
(c) nucleic acid (a) being no more than with above-mentioned nucleic acid a distance in 1000 nucleotide.
5. application according to claim 3 or 4, it is characterised in that: the reagent includes that specific amplification includes methylation
The primer of gene described in claim 1 including site；
Preferably, the diagnostic products are kit, chip or microarray dataset.
6. one group of nucleic acid primer and/or hybridization probe, the potential methylation to one or more gene described in claim 1
Region is specific；
Preferably, the primer and/or hybridization probe are specific in the methylation of amplification marker gene open reading frame
Swim region, especially promoter region；Or methylation is one of following:
(a) nucleic acid defined in the chromosomal foci as shown in claim 1；
(b) comprising the site CpG of nucleic acid a；
(c) nucleic acid (a) being no more than with above-mentioned nucleic acid a distance in 1000 nucleotide.
7. a kind of kit, it is characterised in that: the kit includes that one group of nucleic acid primer as claimed in claim 6 or hybridization are visited
Needle further includes methylation-specific restriction enzyme and/or takes off the reagent of amine for bisulfites nucleotide.
8. a kind of diagnostic products for predicting weightless bone loss bone alkaline phosphatase BALP downside risk, it is characterised in that: described to examine
Stopping pregnancy product include the reagent for being able to detect the level of gene methylation described in claim 1；
Preferably, the reagent of gene methylation level described in the detection claim 1 includes being used for full-length genome methyl
Change screening analysis reagent, for based on chip methylation profiles analysis reagent, for methylation status of PTEN promoter analysis
Reagent, for the reagent of flight mass spectrum detection (sequenome), for the examination of sulphite treated gene order-checking analysis
Agent, the reagent for combining bisulfites restriction endonuclease analysis are used for methylation-specific endonuclease digestion and quantify
The reagent of polymerase chain reaction Conjoint Analysis, for methylation sensitive high-resolution melting curve analysis reagent or for coke
The reagent that phosphoric acid PCR sequencing PCR tests and analyzes.
9. diagnostic products according to claim 8, it is characterised in that: the reagent includes that specific amplification includes methylation
The primer of genetic fragment described in claim 1 including site；
Preferably, the diagnostic products are kit, chip or microarray dataset.
10. the method for predicting weightlessness bone loss bone alkaline phosphatase BALP decline, the method is not with the diagnosing and treating of disease
For the purpose of, it is characterised in that: steps are as follows:
(1) DNA methylation for measuring DNA methylation site is horizontal；
(2) building of two disaggregated model of logistic regression is carried out, and substitutes into following formula:
Wherein, X=intercept coefficient+DNA methylation site DNA methylation level × independent variable coefficient, Y is bone alkaline phosphatase
BALP downside risk score；Such as Y < 0.5, then it is determined as that bone alkaline phosphatase BALP declines.
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|---|---|---|---|---|
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| CN101221171A (en) * | 2007-12-11 | 2008-07-16 | 广州益善生物技术有限公司 | Liquid phase chip used for detecting bone metabolism biochemical marker and its preparing method |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1102708A (en) * | 1993-11-06 | 1995-05-17 | 北京协和医药科技开发总公司 | External diagnosis reagent case for clinicale xamination of baby's bone alkali phosphatase |
| CN101221171A (en) * | 2007-12-11 | 2008-07-16 | 广州益善生物技术有限公司 | Liquid phase chip used for detecting bone metabolism biochemical marker and its preparing method |
| CN203490223U (en) * | 2013-09-03 | 2014-03-19 | 天津朗赛生物科技有限公司 | Joint detection reagent card for ferritin and bone alkaline phosphatase in human blood |
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| Title |
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