CN106701923A - SNP marker relevant with treatment of ischemic stroke through antiplatelet drug clopidogrel and application thereof - Google Patents
SNP marker relevant with treatment of ischemic stroke through antiplatelet drug clopidogrel and application thereof Download PDFInfo
- Publication number
- CN106701923A CN106701923A CN201611103721.0A CN201611103721A CN106701923A CN 106701923 A CN106701923 A CN 106701923A CN 201611103721 A CN201611103721 A CN 201611103721A CN 106701923 A CN106701923 A CN 106701923A
- Authority
- CN
- China
- Prior art keywords
- seq
- clopidogrel
- primer
- snp
- primer sequence
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 239000005552 B01AC04 - Clopidogrel Substances 0.000 title claims abstract description 91
- 229960003009 clopidogrel Drugs 0.000 title claims abstract description 91
- GKTWGGQPFAXNFI-HNNXBMFYSA-N clopidogrel Chemical compound C1([C@H](N2CC=3C=CSC=3CC2)C(=O)OC)=CC=CC=C1Cl GKTWGGQPFAXNFI-HNNXBMFYSA-N 0.000 title claims abstract description 91
- 229940127218 antiplatelet drug Drugs 0.000 title claims abstract description 22
- 238000011282 treatment Methods 0.000 title claims abstract description 10
- 239000003550 marker Substances 0.000 title abstract description 9
- 208000032382 Ischaemic stroke Diseases 0.000 title abstract description 6
- 230000003321 amplification Effects 0.000 claims abstract description 23
- 238000003199 nucleic acid amplification method Methods 0.000 claims abstract description 23
- 239000003814 drug Substances 0.000 claims abstract description 15
- 206010008092 Cerebral artery thrombosis Diseases 0.000 claims description 35
- 208000006011 Stroke Diseases 0.000 claims description 26
- 230000000302 ischemic effect Effects 0.000 claims description 22
- 238000001514 detection method Methods 0.000 claims description 15
- 239000008280 blood Substances 0.000 claims description 9
- 206010008190 Cerebrovascular accident Diseases 0.000 claims description 8
- 102000004190 Enzymes Human genes 0.000 claims description 8
- 108090000790 Enzymes Proteins 0.000 claims description 8
- 230000000857 drug effect Effects 0.000 claims description 8
- 239000003153 chemical reaction reagent Substances 0.000 claims description 7
- 210000005259 peripheral blood Anatomy 0.000 claims description 7
- 239000011886 peripheral blood Substances 0.000 claims description 7
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L magnesium chloride Substances [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 claims description 6
- 238000012360 testing method Methods 0.000 claims description 6
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 6
- 239000008367 deionised water Substances 0.000 claims description 5
- 229910021641 deionized water Inorganic materials 0.000 claims description 5
- 229910001629 magnesium chloride Inorganic materials 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 5
- 230000002490 cerebral effect Effects 0.000 claims description 4
- 230000002068 genetic effect Effects 0.000 claims description 4
- 238000011285 therapeutic regimen Methods 0.000 claims description 3
- 239000013558 reference substance Substances 0.000 claims description 2
- LCCNCVORNKJIRZ-UHFFFAOYSA-N parathion Chemical compound CCOP(=S)(OCC)OC1=CC=C([N+]([O-])=O)C=C1 LCCNCVORNKJIRZ-UHFFFAOYSA-N 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 9
- 229940079593 drug Drugs 0.000 abstract description 8
- 239000000523 sample Substances 0.000 description 12
- 238000003745 diagnosis Methods 0.000 description 10
- 210000004369 blood Anatomy 0.000 description 9
- 238000006243 chemical reaction Methods 0.000 description 9
- 238000000034 method Methods 0.000 description 9
- XTWYTFMLZFPYCI-KQYNXXCUSA-N 5'-adenylphosphoric acid Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](COP(O)(=O)OP(O)(O)=O)[C@@H](O)[C@H]1O XTWYTFMLZFPYCI-KQYNXXCUSA-N 0.000 description 8
- XTWYTFMLZFPYCI-UHFFFAOYSA-N Adenosine diphosphate Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(COP(O)(=O)OP(O)(O)=O)C(O)C1O XTWYTFMLZFPYCI-UHFFFAOYSA-N 0.000 description 8
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 8
- 208000010110 spontaneous platelet aggregation Diseases 0.000 description 8
- 210000001772 blood platelet Anatomy 0.000 description 7
- 201000010099 disease Diseases 0.000 description 7
- 238000003205 genotyping method Methods 0.000 description 7
- 239000003795 chemical substances by application Substances 0.000 description 6
- 239000000203 mixture Substances 0.000 description 6
- 238000011160 research Methods 0.000 description 6
- 238000004458 analytical method Methods 0.000 description 5
- 230000000692 anti-sense effect Effects 0.000 description 5
- 208000026106 cerebrovascular disease Diseases 0.000 description 5
- 238000013461 design Methods 0.000 description 5
- 230000001225 therapeutic effect Effects 0.000 description 5
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- 108010000178 IGF-I-IGFBP-3 complex Proteins 0.000 description 4
- 239000000090 biomarker Substances 0.000 description 4
- PHTQWCKDNZKARW-UHFFFAOYSA-N isoamylol Chemical compound CC(C)CCO PHTQWCKDNZKARW-UHFFFAOYSA-N 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- BSYNRYMUTXBXSQ-UHFFFAOYSA-N Aspirin Chemical compound CC(=O)OC1=CC=CC=C1C(O)=O BSYNRYMUTXBXSQ-UHFFFAOYSA-N 0.000 description 3
- QSMTUAJDOTXEDZ-UHFFFAOYSA-N N1C=CC=C1.[Cl] Chemical compound N1C=CC=C1.[Cl] QSMTUAJDOTXEDZ-UHFFFAOYSA-N 0.000 description 3
- 229960001138 acetylsalicylic acid Drugs 0.000 description 3
- 206010008118 cerebral infarction Diseases 0.000 description 3
- 230000034994 death Effects 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 239000000284 extract Substances 0.000 description 3
- 230000006698 induction Effects 0.000 description 3
- 239000011229 interlayer Substances 0.000 description 3
- 238000007477 logistic regression Methods 0.000 description 3
- 230000035945 sensitivity Effects 0.000 description 3
- 238000004879 turbidimetry Methods 0.000 description 3
- 238000009007 Diagnostic Kit Methods 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 2
- KAESVJOAVNADME-UHFFFAOYSA-N Pyrrole Chemical compound C=1C=CNC=1 KAESVJOAVNADME-UHFFFAOYSA-N 0.000 description 2
- 238000012098 association analyses Methods 0.000 description 2
- 210000004556 brain Anatomy 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 230000002596 correlated effect Effects 0.000 description 2
- 238000002651 drug therapy Methods 0.000 description 2
- 239000007850 fluorescent dye Substances 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000002207 metabolite Substances 0.000 description 2
- 230000000069 prophylactic effect Effects 0.000 description 2
- 239000003223 protective agent Substances 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 230000001629 suppression Effects 0.000 description 2
- 230000009897 systematic effect Effects 0.000 description 2
- OEKWJQXRCDYSHL-FNOIDJSQSA-N ticagrelor Chemical compound C1([C@@H]2C[C@H]2NC=2N=C(N=C3N([C@H]4[C@@H]([C@H](O)[C@@H](OCCO)C4)O)N=NC3=2)SCCC)=CC=C(F)C(F)=C1 OEKWJQXRCDYSHL-FNOIDJSQSA-N 0.000 description 2
- 229960002528 ticagrelor Drugs 0.000 description 2
- 201000001320 Atherosclerosis Diseases 0.000 description 1
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- 108010078791 Carrier Proteins Proteins 0.000 description 1
- 206010013654 Drug abuse Diseases 0.000 description 1
- 208000035150 Hypercholesterolemia Diseases 0.000 description 1
- 206010020772 Hypertension Diseases 0.000 description 1
- 206010061216 Infarction Diseases 0.000 description 1
- 102100025306 Integrin alpha-IIb Human genes 0.000 description 1
- 101710149643 Integrin alpha-IIb Proteins 0.000 description 1
- 102000016267 Leptin Human genes 0.000 description 1
- 108010092277 Leptin Proteins 0.000 description 1
- 206010028851 Necrosis Diseases 0.000 description 1
- 241001597008 Nomeidae Species 0.000 description 1
- 108020005497 Nuclear hormone receptor Proteins 0.000 description 1
- 108700005075 Regulator Genes Proteins 0.000 description 1
- 239000007984 Tris EDTA buffer Substances 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 230000002776 aggregation Effects 0.000 description 1
- 238000004220 aggregation Methods 0.000 description 1
- 239000000556 agonist Substances 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000000903 blocking effect Effects 0.000 description 1
- 210000005013 brain tissue Anatomy 0.000 description 1
- 238000000546 chi-square test Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000000205 computational method Methods 0.000 description 1
- 230000000875 corresponding effect Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000010612 desalination reaction Methods 0.000 description 1
- 206010012601 diabetes mellitus Diseases 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 235000006694 eating habits Nutrition 0.000 description 1
- 230000002526 effect on cardiovascular system Effects 0.000 description 1
- 208000005053 encephalomalacia Diseases 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 229930182470 glycoside Natural products 0.000 description 1
- 150000002338 glycosides Chemical class 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000011337 individualized treatment Methods 0.000 description 1
- 230000007574 infarction Effects 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 239000006101 laboratory sample Substances 0.000 description 1
- NRYBAZVQPHGZNS-ZSOCWYAHSA-N leptin Chemical compound O=C([C@H](CO)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CO)NC(=O)CNC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](N)CC(C)C)CCSC)N1CCC[C@H]1C(=O)NCC(=O)N[C@@H](CS)C(O)=O NRYBAZVQPHGZNS-ZSOCWYAHSA-N 0.000 description 1
- 229940039781 leptin Drugs 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000017074 necrotic cell death Effects 0.000 description 1
- 108020004017 nuclear receptors Proteins 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 238000002205 phenol-chloroform extraction Methods 0.000 description 1
- 230000010118 platelet activation Effects 0.000 description 1
- 239000000106 platelet aggregation inhibitor Substances 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 229940002612 prodrug Drugs 0.000 description 1
- 239000000651 prodrug Substances 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 239000011546 protein dye Substances 0.000 description 1
- 238000013094 purity test Methods 0.000 description 1
- 230000009257 reactivity Effects 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 230000004043 responsiveness Effects 0.000 description 1
- 230000002441 reversible effect Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- FVAUCKIRQBBSSJ-UHFFFAOYSA-M sodium iodide Chemical class [Na+].[I-] FVAUCKIRQBBSSJ-UHFFFAOYSA-M 0.000 description 1
- 235000009518 sodium iodide Nutrition 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 238000000528 statistical test Methods 0.000 description 1
- 239000004575 stone Substances 0.000 description 1
- 230000009861 stroke prevention Effects 0.000 description 1
- 208000011117 substance-related disease Diseases 0.000 description 1
- 229940125670 thienopyridine Drugs 0.000 description 1
- 239000002175 thienopyridine Substances 0.000 description 1
- 229910021642 ultra pure water Inorganic materials 0.000 description 1
- 239000012498 ultrapure water Substances 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 230000002792 vascular Effects 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- 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
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- 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
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- 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
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/106—Pharmacogenomics, i.e. genetic variability in individual responses to drugs and drug metabolism
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- 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
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Wood Science & Technology (AREA)
- Analytical Chemistry (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Pathology (AREA)
- Immunology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses a SNP marker relevant with treatment of ischemic stroke through antiplatelet drug clopidogrel and application thereof, as well as a special amplification primer and a special extension primer for detecting the marker. The SNP marker is combination of rs6686001, rs2302429, rs2242480, rs4244285 and rs2046934. Sequences of each marker, the special amplification primer and the special extension primer are sequentially respectively as shown in SEQ ID No. 1-20. According to the SNP marker combination, clopidogrel-resistant and clopidogrel-effective ischemic stroke people can be distinguished well. The SNP marker, the amplification primer thereof and a relevant kit can be used for clinical medication guidance for adopting the antiplatelet drug clopidogrel to treat ischemic stroke patients, medicine resistance is prevented, and a treatment effect is enhanced.
Description
Technical field
The invention belongs to genetic engineering and cerebrovascular medicine technology field.More particularly, to a kind of and antiplatelet drug
The related SNP marks of thing Effect of Clopidogrel in Treating cerebral arterial thrombosis and its application.
Background technology
Cerebral arterial thrombosis, be otherwise known as cerebral infarction or cerebral infarction（The traditional Chinese medical science is then referred to as apoplexy）, refer to due to brain blood
Liquid supplies obstacle, the ischemic necrosis or cerebromalacia of the limitation brain tissue that ischemic, anoxic cause.In China, as society is sent out
Exhibition and the continuous change of resident living mode and eating habit, cerebral apoplexy have become current China human mortality cause of death second
Disease, seriously threaten residents ' health.
It is narrow in active intervention hypertension, diabetes, hypercholesteremia disease, the cerebrovascular at present according to the evidence of evidence-based medicine EBM
While the hazards of the cerebral arterial thrombosises such as mountain pass, platelet aggregation-against treatment is the foundation stone for effectively preventing and treating cerebral infarction, can be shown
Write the recurrence rate and the death rate for reducing apoplexy.And clopidogrel is anti-after latter important Thienopyridines of aspirin
Antiplatelet drug, it is cardiovascular and cerebrovascular diseases associated to atherosclerosis（Such as miocardial infarction, apoplexy or vascular death）Have
Good preventive and therapeutic effect.Clopidogrel is used as pro-drug, it is necessary to be metabolized by CPY450 enzymes, and generation can suppress platelet aggregation
Active metabolite.The suppression adenosine diphosphate (ADP) of the active metabolite selectivity of clopidogrel（ADP）With its blood platelet P2Y12
The activation of the glycoprotein GPIIb/IIIa compounds of the combination of acceptor and secondary ADP mediations, therefore suppress platelet aggregation.It is logical
Crossing the platelet activation aggregation approach of the ADP inductions of blocking release can also suppress the blood of other agonist inductions in addition to ADP
Platelet is assembled, and reaches good therapeutic effect.
Most of ischemic cerebral stroke patients can be made to obtain good income although taking clopidogrel, got in recent years
More research finds that different patients have different drug responsivenesses to it.Wherein, it was reported that there are about 4~30% patients
Fail to reach the effect for fully suppressing platelet aggregation after taking clopidogrel, but trigger a series of adverse consequences, this phenomenon
It is referred to as clopidogrel Resistant.How clopidogrel Resistant is overcome as asking that numerous clinicians and researcher earnestly pay close attention to
Topic.Numerous inherent causes, such as metabolic enzyme, transporter, nuclear receptor gene polymorphism are considered as having tight association with it.Monokaryon glycosides
Sour polymorphism（SNP）It is considered as imparting individual different phenotypic character, and for factors such as environmental exposure, drug therapies
Differential responses.So if patient has different SNP types on some important related regulatory genes, then can cause difference
Treatment of the patient to clopidogrel has different reactivity, therefore related SNP is probably the key factor of clopidogrel Resistant.
The related SNP acted on by examination clopidogrel is composed, and can be used to screen the beneficiaries of clopidogrel preventing and treating.If can
Combined by related SNP and determine which ischemic cerebral stroke patients is adapted to take clopidogrel, which patient is not suitable for chlorine pyrrole
Gray treats, it becomes possible to determine optimal therapeutic regimen at the initial stage of a disease, reduces drug abuse, reaches good therapeutic effect.
It is clinically urgent problem however, clinically still lack related SNP at present for instructing the rational use of medicines.
The content of the invention
The technical problem to be solved in the present invention is the defect and deficiency for overcoming above-mentioned prior art, there is provided a kind of small with anti-blood
The related SNP site of plate medicine clopidogrel preventing and treating cerebral arterial thrombosis, can be used for using antiplatelet drug Effect of Clopidogrel in Treating
The clinical application of ischemic cerebral stroke patients is instructed, prophylactic agent resistance, strengthens therapeutic effect.Particular by separation and research
SNP in the peripheral blood DNA of the ischemic cerebral stroke patients for taking antiplatelet drug clopidogrel, finds anti-with clopidogrel
Answer the site of sex differernce height correlation, and develop can the clopidogrel of clinical or crowd application prevent and treat the phase of cerebral arterial thrombosis
Kit is closed, for the protective agents selection of cerebral arterial thrombosis provides guidance.
It is an object of the invention to provide a kind of SNP related to antiplatelet drug Effect of Clopidogrel in Treating cerebral arterial thrombosis
Mark.
Another object of the present invention is to provide the specificity amplification primer and specific extension primer of above-mentioned mark.
Another object of the present invention is to provide above-mentioned mark and its specificity amplification primer and exists with specific extension primer
Prepare the application in the auxiliary diagnostic box of antiplatelet drug clopidogrel preventing and treating cerebral arterial thrombosis.
Another object of the present invention is to provide the auxiliary diagnostic box that medication guide is prevented and treated for cerebral arterial thrombosis.
Above-mentioned purpose of the present invention is achieved through the following technical solutions：
A kind of SNP mark related to antiplatelet drug Effect of Clopidogrel in Treating cerebral arterial thrombosis, the SNP marks are
The combination of rs6686001, rs2302429, rs2242480, rs4244285 and rs2046934, its sequence is successively respectively such as SEQ
Shown in ID NO.1~5.The specificity amplification primer of above-mentioned SNP marks is：
The sense primer and downstream primer sequence of rs6686001 specificity amplification primers are respectively such as SEQ ID NO.6 and SEQ ID
NO.7 shows；
The sense primer and downstream primer sequence of rs2302429 specificity amplification primers are respectively such as SEQ ID NO.9 and SEQ ID
Shown in NO.10；
The sense primer and downstream primer sequence of rs2242480 specificity amplification primers are respectively such as SEQ ID NO.12 and SEQ ID
Shown in NO.13；
The sense primer and downstream primer sequence of rs4244285 specificity amplification primers are respectively such as SEQ ID NO.15 and SEQ ID
Shown in NO.16；
The sense primer and downstream primer sequence of rs2046934 specificity amplification primers are respectively such as SEQ ID NO.18 and SEQ ID
Shown in NO.19.
The specific extension primer of above-mentioned SNP marks is：
The specific extension primer sequence of rs6686001 is as shown in SEQ ID NO.8；
The specific extension primer sequence of rs2302429 is as shown in SEQ ID NO.11；
The specific extension primer sequence of rs2242480 is as shown in SEQ ID NO.14；
The specific extension primer sequence of rs4244285 is as shown in SEQ ID NO.17；
The specific extension primer sequence of rs2046934 is as shown in SEQ ID NO.20.
The above-mentioned SNP site of present invention screening, it is related to antiplatelet drug clopidogrel preventing and treating cerebral arterial thrombosis, can use
Instructed in the clinical application using antiplatelet drug Effect of Clopidogrel in Treating ischemic cerebral stroke patients, prophylactic agent resistance increases
Strong therapeutic effect.Particular by separate and research take antiplatelet drug clopidogrel ischemic cerebral stroke patients it is outer
SNP in all blood DNA, find with the site of response to clopidogrel difference height correlation, and develop can clinical or crowd should
Clopidogrel prevents and treats the related kit of cerebral arterial thrombosis, refers to for the protective agents selection of cerebral arterial thrombosis is provided
Lead.
Therefore, it is as described below to apply all be within protection scope of the present invention：
The SNP marks are tried in the preparation auxiliary diagnosis related to antiplatelet drug clopidogrel preventing and treating cerebral arterial thrombosis
Application in agent box.
The specificity amplification primer and/or the specific extension primer are in preparation and antiplatelet drug clopidogrel
Application in the related auxiliary diagnostic box of preventing and treating cerebral arterial thrombosis.
The SNP marks and its specificity amplification primer and/or specific extension primer are preparing guiding treatment ischemic
Application in terms of the preparation or kit of the therapeutic regimen of property brain soldier.
A kind of auxiliary diagnostic box related to clopidogrel preventing and treating cerebral arterial thrombosis, contains detection in the kit
The component of rs6686001, rs2302429, rs2242480, rs4244285 and rs2046934 in peripheral blood DNA.The kit
The SNP of rs6686001, rs2302429, rs2242480, rs4244285 and rs2046934 in peripheral blood DNA is determined for detecting
Mark is combined.
Further, the component includes special described in specificity amplification primer described in claim 2 and/or claim 3
Different in nature extension primer.
Further still, it is preferred that, the component is also including the reagent needed for genetic test.
Preferably, the reagent needed for the genetic test includes Taq enzyme, dNTP mixed liquors, MgCl2Solution, deionized water,
Standard items and/or reference substance etc..
The present inventor gathers standard compliant blood sample, the complete demography money of systematic collection with S.O.P.
Material, clinical data etc., judge whether patient occurs clopidogrel using the platelet aggregation rate of light turbidimetry detection ADP inductions
Resistance, detection of Single locus etc. is carried out using Sequenom MassARRAY Genotypings.Specifically, solve problem of the present invention
Technical scheme include：
（1）The sample storehouse and database of unified standard are set up with S.O.P.（SOP）Standard compliant blood sample is gathered,
The complete demographic data of systematic collection and clinical data.
（2）Genotype detection：Selection ischemic cerebral stroke patients are classified as clopidogrel Resistant group and clopidogrel is non-
Resistance group, in the positive SNP site related with preventing and treating cerebral arterial thrombosis to being found out in the range of clopidogrel related pathways.
（3）The development of the auxiliary diagnostic box of clopidogrel preventing and treating cerebral arterial thrombosis：By patients with cerebral apoplexy according to being
No generation clopidogrel Resistant is layered, and according to the clopidogrel related SNP genotype distribution frequency for detecting, finds out interlayer
SNP with significant difference, develops auxiliary diagnostic box.
The present invention have selected 313 ischemic cerebral stroke patients, wherein resistance group 106, non-resistance group 207.By this
Two groups of crowds obtain correlated results after being detected.
According to the detection of Sequenom MassARRAY, the present inventor detects and compares and obtained in clopidogrel Resistant group
Genotype distribution frequency includes rs6686001, rs2302429 in the SNP that survey interlayer has notable difference and non-resistance group between,
Rs2242480, rs4244285, rs2046934.
Single factor test and multinomial logistic regression result show, the clopidogrel of this 5 SNP and ischemic cerebral stroke patients
Leptin suppression has significantly association.
Further analyze the effect of the combination for Effect of Clopidogrel in Treating cerebral arterial thrombosis auxiliary diagnosis of this 5 SNP, hair
Existing its combination can be good at distinguishing clopidogrel Resistant and the effective cerebral arterial thrombosis crowd of clopidogrel.
According to above-mentioned experimental result, the present inventor is prepared for a kind of clopidogrel that can be used for and prevents and treats the auxiliary of cerebral arterial thrombosis
The kit of diagnosis is helped, comprising the specific primer, the specificity fluorescent probe pair that determine above-mentioned SNP in subject's blood DNA samples
With other detection reagents.
Specifically, the specific primer of this 5 combinations of SNP, or this 5 SNP and specificity fluorescent probe pair
Combining the dependent diagnostic kit for constituting contributes to the auxiliary diagnosis of Effect of Clopidogrel in Treating cerebral arterial thrombosis, is fast clinician
The fast accurate self-condition for grasping patient, takes the control prece of more personalized to provide support.
In addition, present invention also offers a kind of clinic of antiplatelet drug Effect of Clopidogrel in Treating ischemic cerebral stroke patients
Medication guide method, is that the genotype of each SNP is scored, with wild homozygous=" 0 ", other=" 1 ", according to following public affairs
Formula calculates dangerous score value：
Dangerous score value=（0.749 × rs6686001 scores）+（0.810 × rs2302429 scores）-（0.543 ×
Rs2242480 scores）+（0.671 × rs4244285 scores）-（0.714 × rs2046934 scores）；
Subject is assessed according to dangerous score value carries out the preventing and treating of cerebral arterial thrombosis if appropriate for clopidogrel is taken；cutoff
It is 0.2275 to be worth, and dangerous score value is the low-risk crowd of clopidogrel Resistant less than 0.2275, can take clopidogrel preventing and treating
Cerebral arterial thrombosis；And dangerous score value is the High risk group of clopidogrel Resistant more than 0.2275, clopidogrel should not be taken
Preventing and treating cerebral arterial thrombosis.
The invention has the advantages that：
The marker sequence that the present invention is provided changes as the mark of the auxiliary judgment of Effect of Clopidogrel in Treating Ischemic Stroke
It is advantageous in that：
（1）SNP marks are a kind of novel gene biomarkers, it is stable, minimally invasive, be easy to detection, can be examined at the disease initial stage
Go out result, medical diagnosis on disease and treatment-related Sensitivity and Specificity can be effectively improved, the success of such biomarker is opened
Hair will start brand-new situation for the diagnosis and treatment of ischemic cerebral stroke patients, for the development of other diseases biomarker is provided
Use for reference.
（2）Using the appraisement system of scientific system, the present inventor applies Sequenom MassARRAY methods of genotyping
Combined with obtaining the related SNP of disease candidate gene, above method ensure that SNP biomarkers and diagnostic kit in clinic
On application.
（3）The present invention obtain the SNP mark related to the clopidogrel resistance of ischemic cerebral stroke patients and
Specific primer, and carried out the development of related auxiliary diagnostic box.According to detection case, with clinical clopidogrel Resistant
Phenomenon association analysis, has considered the positive and negative association situation and relation intensity of each SNP and clopidogrel Resistant, by danger
Dangerous score value come assess subject if appropriate for take clopidogrel carry out tie Ischemic Stroke prevention and treatment, for height
The patient for resisting risk takes alternative medicine（Such as ticagrelor, aspirin etc.）, clinical application is instructed, improve curative effect of medication.
Brief description of the drawings
Fig. 1 is the ROC curve figure that ischemic cerebral stroke patients occur clopidogrel Resistant.
Specific embodiment
The present invention, but embodiment are further illustrated below in conjunction with Figure of description and specific embodiment not to the present invention
Limit in any form.Unless stated otherwise, reagent, the method and apparatus that the present invention is used are for the art is routinely tried
Agent, method and apparatus.
Unless stated otherwise, following examples agents useful for same and material are purchased in market.
The arrangement of the selection and sample data of the research sample of embodiment 1
1st, samples selection
（1）40~80 years old age, male or female；
（2）Meet the 4th, the whole nation diagnostic criteria of cerebrovascular disease meeting revision, and confirmed through CT or MRI；
（3）With 75mg/ days（Containing being taken with 300mg loading doses first）Oral clopidogrel more than 5 days（Containing 5 days）.
2nd, inventor have selected 313 standard compliant samples and carries out single Sequenom MassARRAY Genotypings altogether
Laboratory sample, and situations such as demographic data and the clinical data of system acquisition these samples.
The detection of the platelet aggregation rate of the sample of embodiment 2 --- the judgement of clopidogrel Resistant
1st, PAgT is carried out using light turbidimetry.Take EDTA anticoagulated whole bloods（1：9）2.7 ml, are centrifuged with 800rpm
5min, extracts Platelet-rich plasm, then 15min is centrifuged with 3000rpm, extracts platelet poor plasma, and platelet count is 10
~20 × 109/ L, PRP to 200~300 × 10 is adjusted with PPP9/ L, blank, 37 DEG C of preheating 3min, with 10 μ are made with PPP
M ADP are derivant, the platelet aggregation rate of light turbidimetry for Determination patient.
2nd, before measure ischemic cerebral stroke patients take clopidogrel（day0）After clopidogrel is taken（day5）Blood
Platelet PAR.If blood platelet inhibiting rate is less than 10% or takes rear blood cluster value after medication>50%, then it is assumed that chlorine pyrrole lattice occur
Thunder is resisted；Patient will be included according to this standard be divided into clopidogrel Resistant group and non-resistance group.
3rd, the change of the platelet aggregation rate before and after clopidogrel is taken by clinical detection patient, 106 diseases have been drawn
There is clopidogrel resistance, 207 patient's non resistances in people.
The parting scanning of SNP marks in the peripheral blood DNA of embodiment 3
In above-mentioned qualified 313 ischemic cerebral stroke patients（Wherein resistance group 106, non-resistance group 207）Through
The detection of Sequenom MassARRAY partings obtains correlated results.Concretely comprise the following steps：
1st, phenol-chloroform method extracts peripheral blood genomic DNA
（1）The μ l of EDTA anticoagulated whole bloods 100 are taken, plus the μ l of aseptic double-distilled water 200 are mixed；
（2）The μ l of 6 M sodium iodides 200 are added, is mixed；
（3）Add chloroform/isoamyl alcohol（24：1, v/v）400 μ l, mix repeatedly, stand 10 min；12000 rpm are centrifuged
10 min, draw supernatant liquor and put another centrifuge tube；
（4）Add the μ l of isopropanol 300 to mix to supernatant, stand 3 min, 12000 rpm are centrifuged 10 min, abandon
Clearly；
（5）The μ l of 70% ethanol 500 of precooling are added, 3 min are stood, 12000 rpm are centrifuged 3 min, discard ethanol；
（6）Repeat step（5）Once, it is inverted a moment；
（7）Sample is put into vacuum desiccator to after being completely dried, and adds the resuspended DNA of μ l of TE buffer solutions 40, treats that DNA fills
After dividing dissolving, the measure of purity and concentration is carried out, preserved with -20 DEG C of conditions.
50~100ng/ μ l DNA are usually led to, purity (ultraviolet 260D and 280D ratios) is 1.6~2.0.
2nd, DNA concentration and purity testing
（1）The μ l of sample 20, plus ultra-pure water are taken to 600 μ l, is fully mixed, determining wavelength with ultraviolet specrophotometer is respectively
OD values when 260 nm, 280 nm, 320 nm, calculate the concentration and purity of DNA.
（2）DNA concentration (μ g/ml)=(OD260-OD320) × 50 μ g/ml × extension rate（30 times of dilution）
（3）DNA purity=(OD260-OD320)/(OD280-OD320)
Standard DNA purity ratio is 1.8；Ratio>1.9, show there is RNA to pollute；Ratio<1.6, show there are the dirts such as protein
Dye, can use chloroform/isoamyl alcohol extracting and purifying again.This DNA used is all qualified.
The Sequenom MassARRAY Genotypings of the SNP of embodiment 4：
1st, take subject's sample, design specific primer, PCR reactions and SNP are determined, detect and compare clopidogrel Resistant group and
The distributional difference of the ischemic cerebral stroke patients different genotype of non-resistance group.
Specific method is as follows：
（1）Design of primers：
Sequenom websites are logged in, is drawn using online Assay Design Suite designs Sequenom IPLEX
（https://seqpws1.sequenom.com/AssayDesignerSuite.html#design:new =
Genotyping）
（2）PCR reaction systems:
DNA array mass spectrum Genotypings PCR system is 7 μ L, including the μ L of DNA 2, deionized water 3.3 μ L, 10 × PCR
Buffer with 20 mM MgCl20.5 μ L, 25 mM MgCl20.4 μ L, 25 mM dNTP Mix 0.1 μ L, 1 μM
Primer Mix 0.5 μ L, 5 U/ μ L PCR Enzyme 0.2 μ L.
（3）PCR reaction conditions such as table 1：
The PCR reaction conditions of table 1
（4）SAP reaction systems：SAP reaction systems are 9 μ L, including 7 μ L PCR primers, deionized water 1.53 μ L, SAP
The μ L of 0.17 μ L, SAP enzyme of buffer 0.3.
（5）SAP reaction conditions such as table 2：
The SAP reaction conditions of table 2
（6）Extension system extension system is 11 μ L, including 9 μ L SAP products, the μ L of deionized water 0.619,
The μ of 0.2 μ L, iPLEX termination mix of iPLEX buffer, 0.2 μ L, Extension primer mix 0.94
The μ L of L, iPLEX enzyme 0.041.
（7）Extension condition such as table 3：
The extension condition of table 3
（8）Chip point sample and data acquisition：After being sample desalination with 15mg CLEAN resins, using MassARRAY RS1000
Reaction solution is transferred on 96-pad Spectro CHIP, operation of setting the chessman on the chessboard according to the chess manual is carried out using MassARRAY Typer work stations, led to
Cross the softwares of Typer 4 and obtain data and collection of illustrative plates.
2nd, data processing and analysis：
Compare the distribution of the target gene frequency in clopidogrel supports resistance group and non-resistance group using logistic regression models
Difference, filters out the 5 positive association SNP that there is significant difference.Specific 5 SNP marks and its amplimer sequence are shown in
Table 4 and table 5.
45 SNP marker sequences of table
The amplimer sequence of the SNP marks of table 5
Note：F：Fowward Primer, upstream primer sequence；R：Reverse Primer, downstream primer sequence；E：Extended
Primer, extension primer sequence.
Embodiment 5 is using MELD method analysis SNP and the relation of clopidogrel Resistant
1st, according to the above results, the present inventor is by carrying out the ratio of genotype distribution frequency to resistance group and non-resistance group sample
Go out 5 positive associations compared with, final choice, with single regression coefficient as weight, further try to achieve dangerous score value, draw ROC and come
The sensitivity and specificity of diagnosis are evaluated, and then analyzes these to taking the situation that antiplatelet drug clopidogrel is resisted
Diagnosed.Conjoint Analysis to 5 marks find, this 5 SNP will be adapted to take clopidogrel and not with 70.9% AUC
The patients with cerebral apoplexy for being adapted to take clopidogrel is separated（As shown in Figure 1）.
2 therefore, inventors demonstrated that use rs6686001, rs2302429, rs2242480, rs4244285,
The combination of rs2046934 can well be distinguished and be adapted to take clopidogrel and be not suitable for taking the crowd of clopidogrel.
That is, the suitable crowd for taking clopidogrel that the combination of this 5 SNP is distinguished is adapted to be entered with this kind of medicine
Row prevention is treated, and being not suitable for of distinguishing takes the crowd of clopidogrel then other antiplatelet drugs of recommended replacement（Such as
Ticagrelor, aspirin etc.）Prevented and treated.
Embodiment 6
It is used for classified variable with Chi-square Test or t is checked and compared demographic characteristics etc. in research object for continuous variable
The difference being distributed between group.Analysis is associated with logistic regression analysis models.
The comprehensive indication constituted to further study this 5 SNP is used to evaluate the effect of auxiliary diagnosis, and we construct
One Mathematical Modeling, considers the relation intensity that each SNP occurs with clopidogrel Resistant.Specifically, we are to each
The genotype of SNP is scored, with wild homozygous=" 0 ", other=" 1 ", logistic recurrence system during with single snp analysis
Number is weight, and the situation for considering each SNP determines a dangerous score value to each research object.
The computational methods of dangerous score value are as follows：
Dangerous score value=（0.749 × rs6686001 scores）+（0.810 × rs2302429 scores）-（0.543 ×
Rs2242480 scores）+（0.671 × rs4244285 scores）-（0.714 × rs2046934 scores）.
The dangerous score value coefficient and boundary value of acquisition are applied directly in 313 samples of association study.According to danger
Dangerous score value carries out the preventing and treating of cerebral arterial thrombosis assessing subject if appropriate for clopidogrel is taken, and cutoff values are
0.2275, and estimate risk accordingly, it is the low-risk crowd of clopidogrel Resistant less than 0.2275 that the dangerous score value of assessment is, can be with
Take clopidogrel and prevent and treat apoplexy.And dangerous score value is the High risk group of clopidogrel Resistant more than 0.2275, it is proposed that use instead
Other medicines are treated.
The case group of table 6 and control group association analysis result
Statistical analysis pass through special statistical analysis software（SPSS 21.0）Complete work.The horizontal P values of significance,statistical
0.05 is set to, all statistical tests are two-sided test.
The making of the related auxiliary diagnostic box of the pyrrole Gray of embodiment 7 preventing and treating cerebral arterial thrombosis
1st, the making of kit and operating process are based on Sequenom MassARRAY genotyping techniques.
After Sequenom MassARRAY are to single detection on candidate gene path, whether occur clopidogrel Resistant this
Under one factor, determine that ischemic cerebral stroke patients genotype distribution frequency interlayer has significant difference, in this, as chlorine pyrrole lattice
Thunder prevents and treats the index of cerebral arterial thrombosis auxiliary diagnosis.What is finally filtered out aids in non-clopidogrel preventing and treating cerebral arterial thrombosis
Diagnose related SNP（Rs6686001, rs2302429, rs2242480, rs4244285, rs2046934）Composition corresponding reagent
Box.
Kit contains primer sequence of a collection of SNP specific primers including following primer（The primer sequence of rs6686001
For the primer sequence that the primer sequence of SEQ ID NO.1-3, rs2302429 is SEQ ID NO.4-6, rs2242480 is SEQ
The primer sequence of ID NO.7-9, rs4244285 is SEQ ID for the primer sequence of SEQ ID NO.10-12, rs2046934
NO.13-15）.
Kit can also be containing the common agents needed for relevant art, such as：DNTPs, MgCl2, distilled water, Taq enzyme etc.,
And for determining the standard items and blank etc. of genotype.
2nd, the value of this kit is to only need to peripheral blood without other tissue samples, by most simplifying and specifically
Primer detection SNP, then composed by SNP and instruct the use of clopidogrel in ischemic cerebral stroke patients, not only stablize, detection side
Just, and accurately, can aid in improving the Sensitivity and Specificity of drug therapy, therefore this kit is put into practice, Ke Yibang
Help guidance diagnosis and more effective individualized treatment.
SEQUENCE LISTING
<110>Zhongshan University, Guangzhou Zhong great Nanshas scientific and technical innovation industrial park Co., Ltd
<120>A kind of SNP mark related to antiplatelet drug Effect of Clopidogrel in Treating cerebral arterial thrombosis and its application
<130>
<160> 20
<170> PatentIn version 3.3
<210> 1
<211> 51
<212> DNA
<213>Mark rs6686001
<400> 1
tgagctgtgc ccaaaggtcc ccaggkgtgg atagtatcca acacatctca a 51
<210> 2
<211> 51
<212> DNA
<213>Mark rs2302429
<400> 2
tggggagggg gcctctgagg tttggrtgcc aggtgggctg gaagaggccc t 51
<210> 3
<211> 51
<212> DNA
<213>Mark rs2242480
<400> 3
cacctcctcc ctccttctcc atgtaycatc cactcacctt attgggtaaa a 51
<210> 4
<211> 51
<212> DNA
<213>Mark rs4244285
<400> 4
ttcccactat cattgattat ttcccrggaa cccataacaa attacttaaa a 51
<210> 5
<211> 51
<212> DNA
<213>Mark rs2046934
<400> 5
tctggtgaaa taaaaagatt acaaaygtca tttcaaattc ccaagatgta g 51
<210> 6
<211> 30
<212> DNA
<213>Rs6686001 sense primers
<400> 6
acgttggatg aatctgagct gtgcccaaag 30
<210> 7
<211> 30
<212> DNA
<213>Rs6686001 anti-sense primers
<400> 7
acgttggatg ctttctggag tgatcctgtg 30
<210> 8
<211> 15
<212> DNA
<213>Rs6686001 extension primers
<400> 8
ccaaaggtcc ccagg 15
<210> 9
<211> 30
<212> DNA
<213>Rs2302429 sense primers
<400> 9
acgttggatg attctcagca tctgtccagc 30
<210> 10
<211> 29
<212> DNA
<213>Rs2302429 anti-sense primers
<400> 10
acgttggatg actcacccag ggcctcttc 29
<210> 11
<211> 16
<212> DNA
<213>Rs2302429 extension primers
<400> 11
ggcctctgag gtttgg 16
<210> 12
<211> 30
<212> DNA
<213>Rs2242480 sense primers
<400> 12
acgttggatg gaagtggtga ggaggcattt 30
<210> 13
<211> 30
<212> DNA
<213>Rs2242480 anti-sense primers
<400> 13
acgttggatg gttttaccca ataaggtgag 30
<210> 14
<211> 17
<212> DNA
<213>Rs2242480 extension primers
<400> 14
ccctccttct ccatgta 17
<210> 15
<211> 31
<212> DNA
<213>Rs4244285 sense primers
<400> 15
acgttggatg gatatgcaat aattttccca c 31
<210> 16
<211> 30
<212> DNA
<213>Rs4244285 anti-sense primers
<400> 16
acgttggatg taaagtcccg agggttgttg 30
<210> 17
<211> 20
<212> DNA
<213>Rs4244285 extension primers
<400> 17
agtaatttgt tatgggttcc 20
<210> 18
<211> 30
<212> DNA
<213>Rs2046934 sense primers
<400> 18
acgttggatg tatggcatct acatcttggg 30
<210> 19
<211> 30
<212> DNA
<213>Rs2046934 anti-sense primers
<400> 19
acgttggatg gaatcaattt cacttatctc 30
<210> 20
<211> 22
<212> DNA
<213>Rs2046934 extension primers
<400> 20
catcttggga atttgaaatg ac 22
Claims (10)
1. a kind of SNP mark related to antiplatelet drug Effect of Clopidogrel in Treating cerebral arterial thrombosis, it is characterised in that institute
State the combination that SNP marks are rs6686001, rs2302429, rs2242480, rs4244285 and rs2046934, its sequence
Successively respectively as shown in SEQ ID NO.1~5.
2. the specificity amplification primer of SNP marks described in a kind of claim 1, it is characterised in that the specific amplification draws
Thing is：
The sense primer and downstream primer sequence of rs6686001 specificity amplification primers are respectively such as SEQ ID NO.6 and SEQ ID
NO.7 shows；
The sense primer and downstream primer sequence of rs2302429 specificity amplification primers are respectively such as SEQ ID NO.9 and SEQ ID
Shown in NO.10；
The sense primer and downstream primer sequence of rs2242480 specificity amplification primers are respectively such as SEQ ID NO.12 and SEQ ID
Shown in NO.13；
The sense primer and downstream primer sequence of rs4244285 specificity amplification primers are respectively such as SEQ ID NO.15 and SEQ ID
Shown in NO.16；
The sense primer and downstream primer sequence of rs2046934 specificity amplification primers are respectively such as SEQ ID NO.18 and SEQ ID
Shown in NO.19.
3. the specific extension primer of SNP marks described in a kind of claim 1, it is characterised in that the specificity extends draws
Thing is：
The specific extension primer sequence of rs6686001 is as shown in SEQ ID NO.8；
The specific extension primer sequence of rs2302429 is as shown in SEQ ID NO.11；
The specific extension primer sequence of rs2242480 is as shown in SEQ ID NO.14；
The specific extension primer sequence of rs4244285 is as shown in SEQ ID NO.17；
The specific extension primer sequence of rs2046934 is as shown in SEQ ID NO.20.
4. SNP marks described in claim 1 are related to antiplatelet drug clopidogrel preventing and treating cerebral arterial thrombosis in preparation
Auxiliary diagnostic box in application.
5. specificity amplification primer described in claim 2 and/or specific extension primer described in claim 3 are being prepared and anti-blood
Application in the related auxiliary diagnostic box of platelet medicine clopidogrel preventing and treating cerebral arterial thrombosis.
6. it is a kind of to prevent and treat the related auxiliary diagnostic box of cerebral arterial thrombosis to clopidogrel, it is characterised in that the kit
In containing detection peripheral blood DNA in rs6686001, rs2302429, rs2242480, rs4244285 and rs2046934 group
Point.
7. kit according to claim 6, it is characterised in that the component includes that specificity expands described in claim 2
Increase specific extension primer described in primer and/or claim 3.
8. kit according to claim 7, it is characterised in that the component also include genetic test needed for reagent.
9. kit according to claim 8, it is characterised in that the reagent needed for the genetic test include Taq enzyme,
DNTP mixed liquors, MgCl2Solution, deionized water, standard items and/or reference substance.
10. special described in specificity amplification primer described in SNP marks, claim 2 described in claim 1 and/or claim 3
Application of the different in nature extension primer in terms of the preparation of therapeutic regimen of guiding treatment ischemic cerebral apoplexy or kit is prepared.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611103721.0A CN106701923B (en) | 2016-12-05 | 2016-12-05 | SNP marker related to treatment of ischemic stroke by antiplatelet drug clopidogrel and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611103721.0A CN106701923B (en) | 2016-12-05 | 2016-12-05 | SNP marker related to treatment of ischemic stroke by antiplatelet drug clopidogrel and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106701923A true CN106701923A (en) | 2017-05-24 |
| CN106701923B CN106701923B (en) | 2020-10-23 |
Family
ID=58935611
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201611103721.0A Active CN106701923B (en) | 2016-12-05 | 2016-12-05 | SNP marker related to treatment of ischemic stroke by antiplatelet drug clopidogrel and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106701923B (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111733221A (en) * | 2019-03-25 | 2020-10-02 | 北京大学第一医院 | ITGA2 gene SNP marker for assisting accurate medication of clopidogrel |
| CN111733225A (en) * | 2019-03-25 | 2020-10-02 | 北京大学第一医院 | PRKG1 gene SNP marker for assisting accurate medication of clopidogrel |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102108387A (en) * | 2009-12-23 | 2011-06-29 | 上海主健生物工程有限公司 | Method for preventing stroke by detecting folate dysbolism |
| RU2014137556A (en) * | 2014-09-17 | 2016-04-10 | Федеральное бюджетное учреждение науки "Центральный научно-исследовательский институт эпидемиологии" Федеральной службы по надзору в сфере защиты прав потребителей и благополучия человека (ФБУН ЦНИИ Эпидемиологии Роспотребнадзора) | The method for determining the individual genetic risk of ischemic stroke |
-
2016
- 2016-12-05 CN CN201611103721.0A patent/CN106701923B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102108387A (en) * | 2009-12-23 | 2011-06-29 | 上海主健生物工程有限公司 | Method for preventing stroke by detecting folate dysbolism |
| RU2014137556A (en) * | 2014-09-17 | 2016-04-10 | Федеральное бюджетное учреждение науки "Центральный научно-исследовательский институт эпидемиологии" Федеральной службы по надзору в сфере защиты прав потребителей и благополучия человека (ФБУН ЦНИИ Эпидемиологии Роспотребнадзора) | The method for determining the individual genetic risk of ischemic stroke |
Non-Patent Citations (4)
| Title |
|---|
| KATHRIN KLEIN等: "Pathway-targeted pharmacogenomics of CYP1A2 in human liver", 《FRONTIERS IN PHARMACOLOGY》 * |
| RUI LIU等: "Associations of CYP3A4, NR1I2, CYP2C19 and P2RY12 polymorphisms with clopidogrel resistance in Chinese patients with ischemic stroke", 《ACTA PHARMACOLOGICA SINICA》 * |
| SEUNG-HUN OH等: "Association between kinase insert domain-containing receptor gene polymorphism and haplotypes and ischemic stroke", 《JOURNAL OF THE NEUROLOGICAL SCIENCES》 * |
| SHU LIU等: "The POR rs1057868–rs2868177 GC-GT diplotype is associated with high tacrolimus concentrations in early post-renal transplant recipients", 《ACTA PHARMACOLOGICA SINICA》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111733221A (en) * | 2019-03-25 | 2020-10-02 | 北京大学第一医院 | ITGA2 gene SNP marker for assisting accurate medication of clopidogrel |
| CN111733225A (en) * | 2019-03-25 | 2020-10-02 | 北京大学第一医院 | PRKG1 gene SNP marker for assisting accurate medication of clopidogrel |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106701923B (en) | 2020-10-23 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Glahn et al. | High dimensional endophenotype ranking in the search for major depression risk genes | |
| CN103957705A (en) | Single nucleotide polymorphisms useful to predict clinical response for glatiramer acetate | |
| Glenn et al. | Significant variance in genetic diversity among populations of Schistosoma haematobium detected using microsatellite DNA loci from a genome-wide database | |
| Williams-Blangero et al. | Host genetics and population structure effects on parasitic disease | |
| CN105177116A (en) | Human CYP2C9 and VKORC1 genetic polymorphism detection kit | |
| JP6188810B2 (en) | Use of the HLA-B * 1301 allele | |
| Afsharpad et al. | Molecular monitoring of Plasmodium falciparum resistance to antimalarial drugs after adoption of sulfadoxine–pyrimethamine plus artesunate as the first line treatment in Iran | |
| CN106701923A (en) | SNP marker relevant with treatment of ischemic stroke through antiplatelet drug clopidogrel and application thereof | |
| CN103911440B (en) | SNP marker related to liver toxicity of platinum type chemotherapeutic medicines and applications thereof | |
| US20210062264A1 (en) | Use of cfdna fragments as biomarkers in patients after organ transplantation | |
| CN104195228B (en) | A kind of SNP mark relevant to nonsyndromic congenital heart disease auxiliary diagnosis and application thereof | |
| CN107557468B (en) | Cancer-testis gene genetic marker related to auxiliary diagnosis of primary lung cancer and application thereof | |
| CN110241197A (en) | Primer combination of probe and kit and application for instructing Atorvastatin drug personalized medicine related gene to detect | |
| Ray et al. | In silico identification of potential drug target and analysis of effective single nucleotide polymorphisms for autism spectrum disorder | |
| CN103602671A (en) | SNP in KRT8 gene exon area and determination method thereof | |
| CN109880903B (en) | SNP marker for auxiliary diagnosis of non-small cell lung cancer and application thereof | |
| CN106868163A (en) | The SNP mark related to the reaction of antituberculotic hepatotoxicity wind agitation, kit and application based on FLT3 and NAT2 genes | |
| CN103290006B (en) | A clinically unexplained NOA-related mitochondrial DNA SNP marker and applications thereof | |
| Ohmori et al. | Development of absolute quantification method for genotype-specific Babesia microti using real-time PCR and practical experimental tips of real-time PCR | |
| Su et al. | Association between the NOTCH4 gene rs3131296 polymorphism with schizophrenia risk in the Chinese Zhuang population and Chinese Han population | |
| CN108753959A (en) | It is a kind of positioned at DISC1FP1 genes and the relevant SNP markers of Brain Radiation Injury caused by radiotherapy and its application | |
| CN107574240A (en) | One NF κ B1 gene insertion/deletion site is preparing the application in detecting hepatitis C neurological susceptibility product | |
| CN106868162A (en) | The SNP mark related to the reaction of antituberculotic hepatotoxicity wind agitation, kit and application based on PPP2R2B and NAT2 genes | |
| CN104263822B (en) | A kind of NOA agnogenic to clinic auxiliary diagnoses relevant low frequency SNV mark and application thereof | |
| Li | Plasmodium falciparum chloroquine resistance marker protein (Pfcrmp) may be a chloroquine target protein in nucleus |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |