CN108070646A - A kind of accurate medication genetic test Solid phase PCR kit of cardiovascular and cerebrovascular disease - Google Patents
A kind of accurate medication genetic test Solid phase PCR kit of cardiovascular and cerebrovascular disease Download PDFInfo
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- 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
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- 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
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- 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
Abstract
The invention discloses a kind of accurate medication genetic test Solid phase PCR kits of cardiovascular and cerebrovascular disease, it is formed including PCR reaction solution mixed liquor and 12 townhouse pipes or 96 hole PCR reaction plates, PCR reaction solution mixed liquor includes primer sets, thermal starting archaeal dna polymerase and PCR reaction buffers.Compared with prior art：1) it is easy to operate：Need to only be added in Solid phase PCR plate it is nucleic acid-templated can directly on machine amplification, and easy to operate, clinical detection specificity is good, and sensitivity is suitable with quantitative PCR, with stronger clinical practice promotional value；2) high sensitivity, high specificity：Solid phase PCR kit disclosed by the invention uses thermal starting archaeal dna polymerase, avoids non-specific amplification, improves detection sensitivity and specificity；3) detection site is complete, is suitable for the guidance of cardiovascular and cerebrovascular disease safe medication；In conclusion the present invention uses the primer sequence of specificity, ensure the reliability of testing result；The detection method is easy to operate, time saving and energy saving；Detection flux is big, and reagent consumables cost is low.
Description
Technical field
The present invention relates to PCR applied technical fields more particularly to a kind of accurate medication gene inspections of cardiovascular and cerebrovascular disease
Survey Solid phase PCR kit.
Background technology
China because the number of cardiovascular and cerebrovascular disease death is up to 3,500,000, just has 1 people to die of cardiovascular and cerebrovascular in average every 10 seconds every year
Disease.As it can be seen that prevention cardiovascular and cerebrovascular disease has become the cardinal task for ensureing human health.Due to everyone genetic cause,
Gender, height and weight, local environment, life style etc. differ, and cause therapeutic effect difference larger.Present clinician uses
Medicine is all to follow treatment guidelines in principle, but guide is directed to group, and individual difference is very big.Accurate medicine passes through base
Because of detection and acquired disposition comprehensive analysis, the therapeutic scheme of individuation is formulated, on the one hand reduces treatment medication and its corresponding
Toxic side effect, on the other hand also can Economy type medicine cost.
In order to avoid Patients with Cardiovascular/Cerebrovascular Diseases injures caused by treating inappropriate medication, its drug susceptibility need to be carried out
Detection, and then realize accurate medication.In this regard, we have chosen the common anti-hypertension of cardiovascular and cerebrovascular disease, arrhythmia cordis, coronary disease
Sick angina pectoris, thrombotic disease, the heart, brain and other arterial circulation disorder diseases, myocardial infarction, apoplexy, venous embolism are anxious
It property and chronic cardiac insufficiency and supraventricular tachycardia, auricular fibrillation and flutters, hyperlipidemia, cardiotropic formulation is examined
It surveys.These drugs include pril (enalapril, lisinopril, benazepil, Perindopril, fosinopril), nitroglycerin,
Warfarin, clopidogrel, aspirin, husky smooth class, Amlodipine, beta-blocker (metoprolol, atenolol, card dimension ground
Lip river), diuretics (Hydrochioro, bumetanide, frusemide, Torasemide, indapamide), Isosorbide Mononitrate, digoxin,
Statins (Simvastatin, Atorvastatin, Pravastatin, Lovastatin), folic acid etc..
Pril drug for hyperpietic there is good targeting protective effect and the prevention of cardiovascular endpoints event to make
With, be particularly suitable for hypertension with after chronic heart failure, myocardial infarction with cardiac insufficiency, diabetic nephropathy, non-saccharide
The sick nephrosis of urine, metabolic syndrome, albuminuria person.The medicine can protect renal function while anti-hypertension, but be strictly twolip
Sword can both improve renal function, it is also possible to cause acute renal failure and hyperkalemia.Nitroglycerin can directly relax vascular smooth
Flesh, particularly thin vessels smooth muscle make systemic vasodilatation, and peripheral resistance is reduced, and venous return is reduced, and are born before and after mitigation heart
Lotus reduces myocardial oxygen consumption, releases myocardial anoxia.For angina pectoris acute attack, acute left ventricular failure is also used for.Warfarin is
Bicoumarin class anticoagulation can hinder thrombosed extension, but without thrombolytic effect.Clopidogrel for prevent and treat because
The heart, brain and other arterial circulation disorder diseases caused by the high aggregation of blood platelet, are platelet aggregation inhibitors, selectively inhibit
The combination of ADP and platelet receptor and the activation for inhibiting the glycoprotein GPIIh/IIIa compounds that ADP is mediated, and inhibit blood platelet
Aggregation.Also platelet aggregation caused by can inhibit non-ad P.Effect to platelet ADP receptor is irreversible.Oral absorption is fast
Speed, protein binding rate is 98% in blood plasma, and in liver metabolism, main metabolites are without antiplatelet aggregative activity.Aspirin
There is inhibitory action to platelet aggregation, the onset risk of acute myocardial infarction AMI suspected patient can be reduced, prevention myocardial infarction recurs,
It is the secondary prevention of apoplexy, reduces the risk of transient ischemic attack and its secondary cerebral apoplexy, can also prevents depth after major operation
Phlebothrombosis and pulmonary embolism reduce cardiovascular risk factors person (familial history of coronary artery disease, diabetes, dyslipidemia, hypertension, fertilizer
Fat, smoking history, the age be more than 50 years old person) myocardial infarction breaking-out risk.Sartans block renin angiotensin-aldehyde
Aldosterone system has significant Renoprotective Effect, in diabetic nephropathy, hypertensive renal damage and Primary Nephrosis patient
In, the effect of sartans is especially prominent, can effectively reduce whole body and glomerulus localised blood pressure, reduces glomerulus to albumen
Permeability, reduce albuminuria, prevent the generation of glomerulosclerosis, delay the development of nephrosis.Amlodipine is suitable for light, moderate
The treatment of high blood pressure, it can also be used to which the treatment of chronic stable angina pectoris can substantially reduce angina pectoris attacking frequency and extension
The patient motion time.Metoprolol, atenolol memory Carvedilol are that clinically the wide oral beta receptor of application hinders at present
Stagnant dose, beta-blocker is as a kind of important cardioactive agents, in diseases such as hypertension, coronary heart disease and heart failure
Drug therapy in occupation of irreplaceable status.Hydrochioro, bumetanide, frusemide, Torasemide, indapamide are
Diuretics arranges sodium by diuresis, reduces volume load, improve the blood pressure increased.Isosorbide Mononitrate is for coronal athero- hard
The property the changed anginal prevention and treatment of heart disease, myocardial infarction is with the anginal anaphase of persistence, pulmonary hypertension, sternly
The cardiac muscle damage of weight.Digoxin is for acute and chronic cardiac insufficiencies such as hypertension, valvular heart disease, congenital heart diseases.
It is particularly suitable for the cardiac insufficiency of the auricular fibrillation with rapid ventricular rate, it may also be used for control the heart with rapid ventricular rate
Atrial fibrillation moves, the ventricular rate of atrial flutter patients and supraventricular tachycardia.Statins is hydroxymethyl glutaryl coenzyme A reductase
Enzyme inhibitor, such drug synthesize rate-limiting enzyme reductase by Reverse transcriptase endogenous cholesterol, block intracellular hydroxyl first penta
Acid metabolic approach reduces intracellular cholesteryl synthesis, so as to feed back sexual stimulus cell membrane surface LDL receptor number
Amount and active increase increase serum cholesterol removing, horizontal reduction.Statins may also suppress liver synthesis apolipoprotein
B-100, so as to reduce the synthesis and secretion rich in triglycerides AV, lipoprotein.Homocysteine is heart disease, the danger of headstroke
One of dangerous factor, folic acid help to decompose homocysteine, reduce cardiopathic occurrence probability.Based on this, it would be desirable to research and develop
A kind of accurate medication genetic test Solid phase PCR kit of cardiovascular and cerebrovascular disease convenient and efficient, that flux is high.
The content of the invention
It is an object of the invention to overcome the deficiencies of the prior art and provide a kind of 13 kinds of heart and brain convenient and efficient, flux is high
The accurate medication gene detecting kit of vascular diseases can detect ACE rs1799752, VKORC1 rs9923231 using it,
CYP2C9*3 rs1057910, NAT2 rs1801279, NAT2 rs1801280, MTRR rs1801394, MTHFR
Rs1801131, MTHFR rs1801133, ADRB1 rs1801253, ADD1 rs4961, NPPA-AS1 rs5065,
CYP2C19*2 rs4244285,CYP2C19*1/17 rs12248560,PON1 rs662,CYP2C19*3 rs4986893,
ALDH2 rs671, CYP2D6 rs1065852, CETP rs708272, GP1BA rs6065, LTC4S rs730012 genes.
The present invention is achieved by the following technical solutions：
A kind of accurate medication genetic test Solid phase PCR kit of cardiovascular and cerebrovascular disease, which is characterized in that reacted including PCR
Liquid mixed liquor and 12 townhouse pipes or 96 hole PCR reaction plates are formed, and the PCR reaction solution mixed liquor includes primer sets, thermal starting DNA
Polymerase and PCR reaction buffers.
Preferably, the primer pair is made of 20 pairs of primers, table 1 specific as follows：
| Detect target | Primer sequence (F/R) |
| ACE rs1799752 | ACCACTCCCATCCTTTCTCC/AGGCTGAGGCAGGAGAATG |
| VKORC1 rs9923231 | CAGGGTTCAAGTGGTTCTCG/AAGGGTAGGTGCAACAGTAAGG |
| CYP2C9*3 rs1057910 | ACGTGTGATTGGCAGAAACC/CCCGGTGATGGTAGAGGTTT |
| NAT2 rs1801279 | CGGGCTGTTCCCTTTGAGA/CACCTGAGGCTGATCCTTCC |
| NAT2 rs1801280 | CGGGCTGTTCCCTTTGAGA/CACCTGAGGCTGATCCTTCC |
| MTRR rs1801394 | GCCTTGAAGTGATGAGGAGG/CCACTGTAACGGCTCTAACC |
| MTHFR rs1801131 | GGCAGAATTTACAGGAATGGC/GGAGGTCTCCCAACTTACCCT |
| MTHFR rs1801133 | CCTCTCCTGACTGTCATCCC/GCCTTCACAAAGCGGAAGAA |
| ADRB1 rs1801253 | TTCAACTGGCTGGGCTACG/GGCCCTACACCTTGGATTCC |
| ADD1 rs4961 | CGGGAGAAGACAAGATGGC/CTGGGTCAGAAAGGCTGGA |
| NPPA-AS1 rs5065 | GGCGAGGAAGTCACCATCA/CCAGGTCACCAAGCCAGAT |
| CYP2C19*2 rs4244285 | AATTACAACCAGAGCTTGGCATA/ACTCCTTGACCTGTTAAACATCC |
| CYP2C19*1/17 rs12248560 | TGGGGCTGTTTTCCTTAGATAA/GCTGGCAGAACTGGGATTT |
| PON1 rs662 | GAAGGCTCCATCCCACATC/AACATACGACCACGCTAAACC |
| CYP2C19*3 rs4986893 | CCTGCAATGTGATCTGCTCC/TGTACTTCAGGGCTTGGTCA |
| ALDH2 rs671 | TACAAGATGTCGGGGAGTGG/TCTCAGGCTTAAAATGGGAAA |
| CYP2D6 rs1065852 | CCATTTGGTAGTGAGGCAGGTA/GGTCCAGCCTGTGGTTTCACCATTT |
| CETP rs708272 | GGCTTTTGAACTTGGCGATAC/GGAGATGGGCTGAGTGGAG |
| GP1BA rs6065 | CCCTGGATCTATCCCACAAT/CCCGTGGAGAAAAGCAAAA |
| LTC4S rs730012 | CGCTGGGACTCCATTCTGA/CACCACTTCCTGCTTGTTCG |
Preferably, the final concentration of 200nM-1uM of the primer, preferably 500nM.
Preferably, the Solid phase PCR kit is that the primer sets, thermal starting archaeal dna polymerase and PCR reaction is slow
Fliud flushing is fixed in 96 hole PCR reaction plates or 12 townhouse PCR reacting holes, and when sample-adding adds nucleic acid-templated progress higher level's amplification.
It is a kind of accurate based on Solid phase PCR kit 13 kinds of cardiovascular and cerebrovascular diseases of detection as described in claim 1-4 is arbitrary
Medication detection method, it is characterised in that：Include the following steps：(1) 13 kind of accurate medication genetic test solid phase of cardiovascular and cerebrovascular disease
The production of PCR amplification kit；(2) buccal swab DNA extraction kit extracts nucleic acid to specifications；(3) Solid phase PCR expands
Reaction；(4) it is sequenced after PCR product electrophoretic analysis.
Preferably, the Solid phase PCR kit described in above-mentioned steps (1) is in the surface-closed containing PCR reaction mixtures
One layer of mineral wax mixture, between 30-38 DEG C, the mineral wax mixture is used to isolate PCR reaction solution the fusing point of mineral wax mixture
In enzyme and other compositions, the mineral wax mixture by atoleine and fusing after solid paraffin mix, the solid
The ratio of paraffin and atoleine is 1:4-1:12, preferably 1:7-1:9.
Preferably, the nucleic acid concentration described in above-mentioned steps (2), not less than 10ng/ul, purity is between 1.8-2.0.
Preferably, the PCR reaction solution mixed liquor is 5 × PCRBuffer (Tris-HClpH8.5 100mM, KCl
500nM、MgCl2 15nM、dNTPs 10mM)、Mg2+, 20 pairs of primers and thermal starting archaeal dna polymerase form.
The present invention provides a kind of accurate medication genetic test Solid phase PCR kits of cardiovascular and cerebrovascular disease, are reacted including PCR
Liquid mixed liquor and 12 townhouse pipes or 96 hole PCR reaction plates are formed, and the PCR reaction solution mixed liquor includes primer sets, thermal starting DNA
Polymerase and PCR reaction buffers.
Compared with prior art, it is 1) easy to operate：Need to only be added in Solid phase PCR plate it is nucleic acid-templated can directly on machine expand
Increase, and it is easy to operate, clinical detection specificity is good, and sensitivity is suitable with quantitative PCR, has stronger clinical practice promotion price
Value；2) high sensitivity, high specificity：Solid phase PCR kit disclosed by the invention uses thermal starting archaeal dna polymerase, avoids non-spy
Specific amplification improves detection sensitivity and specificity；3) detection site is complete, is suitable for the guidance of cardiovascular and cerebrovascular disease safe medication；
In addition, kit disclosed by the invention can disposably detect 20 kinds of target spots relevant with cardiovascular and cerebrovascular disease, cover pril (according to
That Puli, lisinopril, benazepil, Perindopril, fosinopril), nitroglycerin, warfarin, clopidogrel, Ah Si
Woods, husky smooth class, Amlodipine, beta-blocker (metoprolol, atenolol, Carvedilol), diuretics (Hydrochioro, cloth
Mei Tani, frusemide, Torasemide, indapamide), Isosorbide Mononitrate, digoxin, Statins (Simvastatin, Ah cutting down he
Spit of fland, Pravastatin, Lovastatin), 13 kinds of related drugs such as folic acid.
In conclusion the present invention provides a kind of accurate medication genetic test Solid phase PCR kit of cardiovascular and cerebrovascular disease, this hair
The bright primer sequence using specificity ensures the reliability of testing result；The detection method is easy to operate, time saving and energy saving；Detection
Flux is big, and reagent consumables cost is low；It can be directly to the detection of nucleic acids of extraction；Requirement to detection platform and testing staff is low, energy
It is enough to be widely popularized in a clinical line.
Description of the drawings
Fig. 1 is 12 townhouse figure of kit；
Fig. 2 detects 1 electrophoretogram of patient for kit；
Fig. 3 detects 1 sequencing result of patient for kit.
Specific embodiment
It elaborates below to the embodiment of the present invention, the present embodiment is carried out lower based on the technical solution of the present invention
Implement, give detailed embodiment and specific operating process, but protection scope of the present invention is not limited to following implementation
Example.
The composition of the accurate medication Solid phase PCR detection kit of 1 13 kinds of cardiovascular and cerebrovascular diseases of embodiment
The kit is made of PCR reaction solution mixed liquor and 12 townhouse pipes or 96 hole PCR reaction plates, and wherein PCR reactions are mixed
Conjunction liquid is 5 × PCRBuffer (Tris-HCl pH8.5 100mM, KCl 500nM, MgCl2 15nM、dNTPs10mM)、Mg2+、
20 pairs of primers and thermal starting archaeal dna polymerase are formed.
The kit is one layer of mineral wax mixture of surface-closed in above-mentioned PCR reaction mixtures, and mineral wax mixture is by solid
Body paraffin and atoleine are according to 1:8 (W/V) are mixed.
The reaction system (25ul) of the kit is：PCR reaction solution 23ul, template 2ul.
The operation of 2 kit of embodiment and result judgement
1st, the extraction of genomic DNA
Using buccal swab extract population inner cavity surface cast-off cells, with buccal swab genome DNA extracting reagent kit (my god
Root biochemical technology, article No.：DP322) DNA is extracted according to kit specification.
2nd, the preparation of reaction system
The accurate 12 townhouse pipe of medication genetic test Solid phase PCR reaction kit of cranial vascular disease of coring or 96 hole PCR are anti-
Plate is answered, adds in the DNA of 2ul extractions.
3rd, PCR amplification
PCR pipe is put into regular-PCR instrument and suggests that just doing program carries out PCR amplification to specifications.
4th, 1.0% agarose gel electrophoresis analysis PCR product
5th, result interpretation
This kit results interpretation standard such as the following table 1
| Detect target | Amplified fragments size (bp) |
| ACE rs1799752 | 174 |
| VKORC1 rs9923231 | 278 |
| CYP2C9*3 rs1057910 | 253 |
| NAT2 rs1801279 | 342 |
| NAT2 rs1801280 | 342 |
| MTRR rs1801394 | 159 |
| MTHFR rs1801131 | 226 |
| MTHFR rs1801133 | 168 |
| ADRB1 rs1801253 | 355 |
| ADD1 rs4961 | 279 |
| NPPA-AS1 rs5065 | 248 |
| CYP2C19*2 rs4244285 | 356 |
| CYP2C19*1/17 rs12248560 | 250 |
| PON1 rs662 | 234 |
| CYP2C19*3 rs4986893 | 214 |
| ALDH2 rs671 | 345 |
| CYP2D6 rs1065852 | 292 |
| CETP rs708272 | 416 |
| GP1BA rs6065 | 371 |
| LTC4S rs730012 | 295 |
3 sequencing result of embodiment is understood and medication guide
Using the DNA of 1 Patients with Cardiovascular/Cerebrovascular Diseases buccal swab of embodiment extraction as template, after being expanded with above-mentioned kit,
Following sequencing result is obtained, it is as follows to provide corresponding medication guide according to sequencing result：
The foregoing is merely illustrative of the preferred embodiments of the present invention, is not intended to limit the invention, all essences in the present invention
All any modification, equivalent and improvement made within refreshing and principle etc., should all be included in the protection scope of the present invention.
Claims (8)
1. a kind of accurate medication genetic test Solid phase PCR kit of cardiovascular and cerebrovascular disease, which is characterized in that including PCR reaction solution
Mixed liquor and 12 townhouse pipes or 96 hole PCR reaction plates are formed, and the PCR reaction solution mixed liquor includes primer sets, thermal starting DNA gathers
Synthase and PCR reaction buffers.
2. kit according to claim 1, it is characterised in that：The primer pair is made of 20 pairs of primers, specifically such as
Under：
It is described detection ACE rs1799752 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.1,
Shown in SEQIDNO.2；
It is described detection VKORC1 rs9923231 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.3,
Shown in SEQIDNO.4；
It is described detection CYP2C9*3 rs1057910 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.5,
Shown in SEQIDNO.6；
It is described detection NAT2 rs1801279 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.7,
Shown in SEQIDNO.8；
It is described detection NAT2 rs1801280 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.9,
Shown in SEQIDNO.10；
It is described detection MTRR rs1801394 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.11,
Shown in SEQIDNO.12；
It is described detection MTHFR rs1801131 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.13,
Shown in SEQIDNO.14；
The nucleotide sequence SEQIDNO.15 of the upstream and downstream primer of the detection MTHFR rs1801133 primer pairs,
Shown in SEQIDNO.16；
The nucleotide sequence SEQIDNO.17 of the upstream and downstream primer of the detection ADRB1 rs1801253 primer pairs,
Shown in SEQIDNO.18；
It is described detection ADD1 rs4961 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.19,
Shown in SEQIDNO.20；
It is described detection NPPA-AS1 rs5065 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.21,
Shown in SEQIDNO.22；
It is described detection CYP2C19*2 rs4244285 primer pairs upstream and downstream primer nucleotide sequence respectively by
Shown in SEQIDNO.23, SEQIDNO.24；
It is described detection CYP2C19*1/17 rs12248560 primer pairs upstream and downstream primer nucleotide sequence respectively by
Shown in SEQIDNO.25, SEQIDNO.26；
It is described detection PON1 rs662 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.27,
Shown in SEQIDNO.28；
It is described detection CYP2C19*3 rs4986893 primer pairs upstream and downstream primer nucleotide sequence respectively by
Shown in SEQIDNO.29, SEQIDNO.30；
It is described detection ALDH2 rs671 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.31,
Shown in SEQIDNO.32；
It is described detection CYP2D6 rs1065852 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.33,
Shown in SEQIDNO.34；
It is described detection CETP rs708272 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.35,
Shown in SEQIDNO.36；
It is described detection GP1BA rs6065 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.37,
Shown in SEQIDNO.38；
It is described detection LTC4S rs730012 primer pairs upstream and downstream primer nucleotide sequence respectively by SEQIDNO.39,
Shown in SEQIDNO.40.
3. kit according to claim 1, it is characterised in that：The final concentration of 200nM-1uM of the primer, preferably
500nM。
4. kit according to claim 1, it is characterised in that：The Solid phase PCR kit is by the primer
Group, thermal starting archaeal dna polymerase and PCR reaction buffers are fixed in 96 hole PCR reaction plates or 12 townhouse PCR reacting holes, are added
Nucleic acid-templated progress higher level's amplification is added during sample.
5. a kind of precisely being used based on Solid phase PCR kit 13 kinds of cardiovascular and cerebrovascular diseases of detection as described in claim 1-4 is arbitrary
Medicine detection method, it is characterised in that：Include the following steps：(1) 13 kind of accurate medication genetic test Solid phase PCR of cardiovascular and cerebrovascular disease
The production of amplification kit；(2) buccal swab DNA extraction kit extracts nucleic acid to specifications；(3) Solid phase PCR amplification is anti-
It should；(4) it is sequenced after PCR product electrophoretic analysis.
6. a kind of Solid phase PCR kit 13 kinds of cardiovascular and cerebrovascular diseases of detection that are based on according to claim 5 precisely use medicine inspection
Survey method, it is characterised in that：Solid phase PCR kit described in above-mentioned steps (1) is in the surface envelope containing PCR reaction mixtures
One layer of mineral wax mixture is closed, between 30-38 DEG C, the mineral wax mixture reacts the fusing point of mineral wax mixture for isolating PCR
Enzyme and other compositions in liquid, the mineral wax mixture is mixed by the solid paraffin after atoleine and fusing, described solid
The ratio of body paraffin and atoleine is 1:4-1:12, preferably 1:7-1:9.
7. a kind of Solid phase PCR kit 13 kinds of cardiovascular and cerebrovascular diseases of detection that are based on according to claim 5 precisely use medicine inspection
Survey method, it is characterised in that：Nucleic acid concentration described in above-mentioned steps (2) is not less than 10ng/ul, and purity is between 1.8-2.0.
8. according to the arbitrary kit of claim 1 or 6, it is characterised in that：The PCR reaction solution mixed liquor for 5 ×
PCRBuffer(Tris-HCl pH8.5 100mM、KCl 500nM、MgCl2 15nM、dNTPs 10mM)、Mg2+, 20 pairs of primers
And thermal starting archaeal dna polymerase is formed.
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| CN201710485408.6A CN108070646A (en) | 2017-06-23 | 2017-06-23 | A kind of accurate medication genetic test Solid phase PCR kit of cardiovascular and cerebrovascular disease |
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| CN109022430A (en) * | 2018-07-28 | 2018-12-18 | 山东德诺生物科技有限公司 | For detecting the primed probe group and its application of rs730012 |
| CN109371019A (en) * | 2018-08-23 | 2019-02-22 | 山东德诺生物科技有限公司 | For detecting the primed probe group and its application of rs6065 |
| CN109402251A (en) * | 2018-12-14 | 2019-03-01 | 武汉康昕瑞基因健康科技有限公司 | Cardiovascular and cerebrovascular diseases medicament related gene parting detection primer group, kit and detection method |
| CN110229881A (en) * | 2019-07-03 | 2019-09-13 | 贵州省临床检验中心 | One kind SNP marker relevant to adult cardiovascular disease and its application |
| CN110423803A (en) * | 2019-08-07 | 2019-11-08 | 郑州大学第一附属医院 | Clopidogrel, statin and aspirin relevant drug metabolism Genotyping detection composite amplification system and kit |
| CN111593100A (en) * | 2019-02-20 | 2020-08-28 | 葛猛 | Artificial simulated nucleic acid molecular beacon and kit for detecting GP1BA gene rs6065 site polymorphism |
| CN112080563A (en) * | 2020-09-01 | 2020-12-15 | 美思睿(北京)生物科技有限公司 | Kit for detecting accurate medication genes of chronic diseases |
| CN112553319A (en) * | 2020-12-10 | 2021-03-26 | 北京大学人民医院 | Special primer for detecting anticoagulant and antiplatelet drug resistance related gene SNP locus and application |
| CN113088572A (en) * | 2021-05-25 | 2021-07-09 | 杭州方略生物科技有限公司 | Solid phase PCR kit for hepatitis B drug detection |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101560555A (en) * | 2009-03-18 | 2009-10-21 | 上海中优医药高科技有限公司 | Gene detection method used for individualistic education and health guidance of children |
| CN105018583A (en) * | 2014-04-28 | 2015-11-04 | 博奥生物集团有限公司 | Kit used for detecting polymorphism of genes related to Warfarin and Clopidogrel personalized medication and application thereof |
| CN106367479A (en) * | 2016-08-25 | 2017-02-01 | 杭州百迈生物股份有限公司 | Detection composition for guiding hypertension medication, applications of detection composition, kit and detection method |
-
2017
- 2017-06-23 CN CN201710485408.6A patent/CN108070646A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101560555A (en) * | 2009-03-18 | 2009-10-21 | 上海中优医药高科技有限公司 | Gene detection method used for individualistic education and health guidance of children |
| CN105018583A (en) * | 2014-04-28 | 2015-11-04 | 博奥生物集团有限公司 | Kit used for detecting polymorphism of genes related to Warfarin and Clopidogrel personalized medication and application thereof |
| CN106367479A (en) * | 2016-08-25 | 2017-02-01 | 杭州百迈生物股份有限公司 | Detection composition for guiding hypertension medication, applications of detection composition, kit and detection method |
Non-Patent Citations (5)
| Title |
|---|
| 刘芳 等: ""NAT2和CYP2E1基因多态性及其代谢表型在中国汉族儿童中的分布"", 《中国当代儿科杂志》 * |
| 李娜 等: "血管紧张素转换酶基因rs1799752位点插入或缺失多态性与糖尿病视网膜病变的相关性研究"", 《中华眼科杂志》 * |
| 杨琳 等: ""基于620份全外显子组数据的阿司匹林药物基因组学分析"", 《中华儿科杂志》 * |
| 瞿贤和 等: ""CETP基因rs708272与肥胖青少年运动调节血脂效果的关联性及分子调控机制"", 《武汉体育学院学报》 * |
| 龚平原 等: ""钠离子转运相关基因多态与原发性高血压的相关性研究"", 《中华医学遗传学杂志》 * |
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| CN112080563A (en) * | 2020-09-01 | 2020-12-15 | 美思睿(北京)生物科技有限公司 | Kit for detecting accurate medication genes of chronic diseases |
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