CN105586406A - Method for detecting gene polymorphism of ADRB1 and GRK5 - Google Patents
Method for detecting gene polymorphism of ADRB1 and GRK5 Download PDFInfo
- Publication number
- CN105586406A CN105586406A CN201610029394.2A CN201610029394A CN105586406A CN 105586406 A CN105586406 A CN 105586406A CN 201610029394 A CN201610029394 A CN 201610029394A CN 105586406 A CN105586406 A CN 105586406A
- Authority
- CN
- China
- Prior art keywords
- reagent
- adrb1
- grk5
- sequence
- arbitrary
- 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.)
- Pending
Links
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 provides a genetic marker for prognosis and/or beta receptor inhibitor drug treatment effect evaluation of a patient with heart failure, a reagent for detecting the genetic marker and a kit. The genetic marker is used for gene typing of ADRB1-Arg389Gly (rs1801253) and/or GRK5-Gln41Leu(rs17098707) polymorphic sites. The kit can be used for detecting the gene typing of ADRB1-Arg389Gly (rs1801253) and/or GRK5-Gln41Leu(rs17098707) polymorphic sites of the patient, and then guidance is provided for anticipation of prognosis and/or beta receptor inhibitor drug treatment effect of the patient with heart failure.
Description
Technical field:
The invention belongs to life science, relate to medical science and biotechnology, specifically provide different detection methods (including but not limited to improvementThe method of sequence measurement and Taqman probe Genotyping) simultaneously to beta receptor signal path related gene ADRB1, on GRK5 2Genotyping is fast and accurately carried out in SNP site (ADRB1:rs1801253 and GRK5:rs17098707), is convenient to clinical β is subject toThe curative effect of body retarding agent is assessed.
Background technology:
In heart failure (abbreviation heart failure) is a kind of not enough and cause many organ perfusions of whole body deficiency and circulation extravasated blood as performance taking cardiac pumpingComplex clinical syndrome, is associated with the abnormal change of neuroendocrine system, is the multiple heart including hypertension, coronary heart disease, cardiomyopathyThe end stage eventually of vascular diseases. Beta-3 adrenergic receptor (abbreviation beta receptor) retarding agent is because being by the overactive adrenal gland of blocking-upSystem, effectively reduces heart oxygen consumption, improves myocardial remodelling and patient's long-term prognosis, is used for the multiple painstaking effort including heart failure as standard schemeThe treatment of pipe disease. But more clinical testing and polycentric meta analyze the overall benefit of report beta-blocker from part population,Show individual difference and racial diversify.
At present, the gene pleiomorphism of existing numerous bibliographical information Beta-3 adrenergic receptor paths can affect the curative effect of beta-blocker, and thenThe potential prognosis that affects patients with heart failure. 1040 people's that the people such as StephenB.Liggett deliver for 2006 European heart failure crowd'sResearch shows that β1receptor Gene A DRB1-Arg389Gly position polymorphism causes amino acid change to affect the result for the treatment of of bucindolol. With peaceConsole agent group and compare, the mortality that the patient of ADRB1-Arg-389 homozygous genotype accepts the treatment of beta-blocker bucindolol declines38% (p=0.03), then admission rate 34% (p=0.004) that decline, and the genotype genotypic patient treatment group that is ADRB1-Gly-389 andPrognosis no difference of science of statistics between placebo. The improvement that another 224 routine patients with heart failure research is treated rear heart function by detection has confirmedThe impact of ADRB1-389 site on beta-blocker result for the treatment of, is specially: compared with the genotypic patient of ADRB1-Gly-389,After the patient treatment of ADRB1-Arg389-homozygous genotype, Left Ventricular Ejection Fraction obviously improves (8.7 ± 1.1%vs.0.93 ± 1.7%, p < 0.02).GRK5 is the g protein coupled receptor kinases in beta receptor downstream, in the polymorphism GRK5Gln41Leu site of its 41 amino acids in AfricaCrowd's less important gene frequency is approximately 10 times of European crowd. Two researchs prove in the situation that not taking beta-blocker simultaneouslyThe patient's of GRK5-Leu41 long term survival rate higher (Logrankp=0.013). And the genotypic patient of GRK5-Gln41 is to beta receptorThe therapeutic response of retarding agent is better, is embodied in the life span of non-heart transplant after the patient treatment of GRK5-Gln41 homozygous genotype moreLong (HR=0.22; 95%CI, 0.12-0.40; P < 0.001), and whether the genotypic patient of GRK5-Leu41 accepts beta-blocker controlTreat prognosis no difference of science of statistics (HR=0.78; 95%CI, 0.35-1.17; P=0.53).
The gene pleiomorphism that we sum up beta adrenergic system from existing research does not play a major role in the generation of heart failure, but energyThe clinical phenotypes that modified beta ARBs is relevant, comprises the improvement of beta-blocker on short-term heart function and the impact of long-term survival rate, rightClinical guidance Personalized medicine and result for the treatment of assessment have great importance. But to so far, according to bibliographical information, for beta receptor kidneyThe method that the detection method of upper parathyrine energy system gene polymorphism is direct Sequencing comprises traditional Sanger PCR sequencing PCR and Pyrosequencing order-checkingMethod. Its advantage is that sequencing result is accurate, but operating process is more complicated, and consuming time longer, cost is higher, has limited it in actual clinical workDevelopment in work and application, cannot meet the demand that instructs fast clinical assessment, a kind of new fast, prepare, detection method becomes cheaplyFor active demand.
Summary of the invention:
For making up the deficiencies in the prior art, the present invention aim to provide for above ADRB1-Arg389Gly (rs1801253) and/or2 pleomorphism sites of GRK5-Gln42Leu (rs17098707) detection method fast and accurately, includes but not limited to improvement order-checking and TaqmanThe method of probe Genotyping, for the expected effect of the prognosis of clinical assessment patients with heart failure and/or beta-blocker provides guidance program.
For this reason, the invention provides following technical scheme:
The invention provides the genetic marker of a kind of patients with heart failure prognosis and/or beta-blocker medicine treatment curative effect evaluation, wherein, described something lostPassing mark is the gene of ADRB1-Arg389Gly (rs1801253) and/or GRK5-Gln41Leu (rs17098707) pleomorphism siteSomatotype. Preferably, described genetic marker is (1) ADRB1-Arg389Gly (rs1801253) or (2) GRK5-Gln41Leu(rs17098707) Genotyping of pleomorphism site.
The present invention also provides some reagent for detection of above-mentioned mark.
Preferably, described reagent comprise 1, for the specificity amplification primer pair that comprises above-mentioned 2 corresponding target sequences in SNP site; With/ or 2, identification pleomorphism site detector probe.
Preferred, described for the right structure of the specificity amplification primer of above 2 corresponding target sequences in SNP site as SEQIDNo.Shown in 1-4 or as SEQINNo.5-6 or SEQIDNo.9-10.
The primer sequence SEQIDNo.1-2 of the object fragment that preferably, comprises rs1801253 site for increasing; Be used for the rs17098707 that increasesThe primer SEQIDNo.3-4 of the object fragment in site. Or comprise the object fragment primer sequence SEQID in rs1801253 site for increasingNo.5-6; For the object fragment primer sequence SEQINNo.9-10 that increases and comprise rs17098707 site.
The detector probe of described identification pleomorphism site is the oligonucleotide sequence of 14-50 nucleic acid, can specific and above-mentioned pleomorphism siteSpecific hybrid. Further, described identification probe sequence by 5 ' end fluorescence report group, 3 ' end non-fluorescent quenching group and with polymorphicProperty site before and after the sequence composition of sequence hybridization. Preferred, described fluorescence report group is Fam, Vic etc., and 3 ' the non-fluorescence of end is suddenThe group that goes out is MGB modification group.
Preferred, the described detector probe structure for above 2 SNP sites is as SEQIDNo.7-8 or SEQIDNo.11-12Shown in.
Preferably, for identifying the probe sequence SEQIDNo.7-8 of rs1801253 site allele A/G; Be used for identifying rs17098707The probe sequence SEQIDNo.11-12 of site allele C/G.
The present invention also provides the kit of a kind of mentioned reagent in preparation assessment patients with heart failure prognosis and/or beta-blocker medicine treatment curative effectIn purposes.
The present invention also provides a kind of kit for assessment of patients with heart failure prognosis and/or beta-blocker medicine treatment curative effect, described kitComprise the reagent that at least one is selected from above-mentioned any one. Preferably, described kit also comprises the suitable buffer system for detection of reactionAnd detection architecture.
Preferably, described beta-blocker includes but are not limited to metoprolol, bisoprolol, bucindolol, Carvedilol.
The present invention also provides the method that detects above-mentioned genetic marker in clinical samples, wherein, in described method, utilizes mentioned reagent.
Preferably, described clinical samples is selected from: PBC, leucocyte, serum, urine sample, saliva, body fluid and/or (biopsy) tissueSample, preferred, described sample carries out purifying in advance, for example, separate total DNA.
Preferably, described method comprises the method for (1) improvement order-checking; (2) method of Taqman probe Genotyping; And/or (3) itsHe is based on specific amplification and/or identify above pleomorphism site and the method for sequence around.
Preferred, in the method for described (1) improvement order-checking, comprise and using for comprising the special of above-mentioned SNP site respective objects sequenceProperty amplimer.
Further, in the method for described (1) improvement order-checking, the described specificity for comprising the corresponding target sequence in above-mentioned SNP site expandsIncrease primer pair structure, as SEQIDNo.1-4.
Preferred, in described (2) Taqman probe genotyping detection method, comprise and using for comprising the corresponding target in above-mentioned SNP siteThe detector probe of the specificity amplification primer of sequence and above-mentioned identification pleomorphism site detects genetic marker.
Further, in described (2) Taqman probe Genotyping detection method, described for comprising the corresponding target order in above-mentioned SNP siteThe right structure of specificity amplification primer of row is as shown in SEQIDNo.5-6 or SEQINNo.9-10. The spy of described identification pleomorphism siteNeedle construction is as SEQIDNo.7-8 or SEQIDNo.11-12.
Further, described (3) other based on specific amplification and/or identify above pleomorphism site and around the method for sequence also comprise high scoreDistinguish the method, liquid phase DNA chip typing method, solid phase DNA chip typing method of rate solubility curve, for the PCR that comprises object fragmentThe analytical technique of mass spectrum of product, restriction enzyme typing method, allele-specific hybridization technique, oligonucleotides connect experiment etc.
The invention has the advantages that:
1. the detection method of the order-checking of the improvement for ADRB1rs1801253 and GRK5rs17098707 site provided by the invention is surveyed in traditionOn the basis of order method, optimize design of primers, pcr amplification primer can be used as sequencing primer simultaneously, has reduced design of primers cost, has simplified behaviourDo.
2. method optimizing reaction system on the basis of traditional sequence measurement of improvement order-checking provided by the invention, before the correct accuracy of guaranteePut the consumption that has greatly reduced sequencing reagent, can be reduced to 1/16 of standard consumption, greatly reduce testing cost.
3. method Optimum Operation flow process on the basis of traditional sequence measurement of improvement provided by the invention order-checking, in 1 day from sample process to obtainingTesting result, has simplified operating procedure greatly, has realized fast detecting, meets the needs that clinical reagent uses.
4. the method for Taqman probe somatotype provided by the invention is made optimization on primer and probe sequence design and experimental program, is protectingUnder the prerequisite of card experimental result accuracy, adopt the method for partly measuring reagent, effectively saved testing cost, can better meet clinical practiceDemand. As preferably 3 ' the non-fluorescent quenching group of end is MGB modification group, ensures the specificity and the success rate that detect.
Brief description of the drawings
Fig. 1. utilize the method for improvement order-checking ADRB1rs1801253Arg389Gly site to be carried out to the diagram of Genotyping
Fig. 1-top represents that offside complementary even allele in ADRB1rs1801253 site is G, and patient's genotype is CC type, and Fig. 1-middle part representsADRB1rs1801253 loci gene type is CG heterozygous, and Fig. 1-bottom represents ADRB1rs1801253 site offside complementary strand equipotential baseBecause CC, patient's genotype is GG type.
Fig. 2. utilize the method for improvement order-checking GRK5rs17098707 site to be carried out to the diagram of Genotyping
Fig. 2-top represents that GRK5rs17098707 loci gene type is AA type, and Fig. 2-bottom represents that GRK5rs17098707 loci gene type isAT type.
Fig. 3. utilize the method for Taqman probe ADRB1rs1801253Arg389Gly site to be carried out to the diagram of Genotyping
In the loose some representative sample of bottom right, Fam fluorescence only detected, experimenter's genotype is GG (bottom right). The loose point in upper left represents only to detect in sampleVic fluorescence, experimenter's genotype is CC (upper left). In middle loose some representative sample, Fam fluorescence both detected, Vic fluorescence detected again,Experimenter's genotype is heterozygous type CG (centre).
Fig. 4. utilize the method for Taqman probe GRK5rs17098707 site to be carried out to the diagram of Genotyping
In the loose some expression in bottom right sample, Fam fluorescence only detected, experimenter's genotype is wild type AA (bottom right). In in upper loose some representative sample bothFam fluorescence detected, Vic fluorescence detected again, experimenter's genotype is that wild sudden change mixes genotype AT (on).
Detailed description of the invention:
Below in conjunction with specific embodiment, technical scheme of the present invention is further described. Be understandable that implementation-specific described hereThe mode of executing represents by way of example, and it is not as limitation of the present invention. In the situation that not deviating from the scope of the invention, thisBright principal character can be for various embodiments. One skilled in the art will appreciate that maybe and can confirm, only use normal experiment,Many equivalents can be applied in particular step described herein. These equivalent places of being considered within the scope of the present invention, and quiltClaim covers.
The method of embodiment 1 improvement order-checking is determined the genotype of ADRB1rs1801253 and rs17098707 pleomorphism site
We have been selected in the Patients with Cardiac Failure that 50 routine Wuhan Tongji University heart internal medicine in hospitals are in hospital, and patient's inclusion criteria is: 1,18-80 year is because of heart failureThe patient who is in hospital; 2, assess and determine that New York Heart Association is in II-IV level through at least three clinicians; 3, shrinkage cardiac insufficiencyOr diastolic cardiac insufficiency is all included. 4, include patient in and all pass through perfect physical examination, blood test, electrocardiogram and iconographyCheck.
(1) peripheral blood sample DNA extracts: all experimenters, in the situation of not taking medicine of being admitted to hospital, use EDTA-K2The heparin tube of anti-freezing is adoptedCollection peripheric venous blood 5ml. With 3000 turn/min centrifugal 8 minutes, separate leukocytic cream. Use poba gene group DNA to extract kit(TIANGENBIOTECH[BEIJING] GO., Ltd) extraction leucocyte DNA.
(2) determine the genotype of ADRB1rs1801253 and rs17098707 pleomorphism site by the method for improvement order-checking, concrete schemeFor:
Required primer sequence is synthesized by Hua Da bio tech ltd, after primer sequence is synthetic, is dissolved as 100Um/L with Tris-EDTA solutionConcentration preserve. Other main agents be PCR reaction system (comprise thermal starting enzyme, 2*GCbuffer, 2.5MdNTP, is purchased from ShanghaiThe precious biotech firm in Dalian), sequencing reaction system (HIDI, is purchased from American AB I company for 2.5*bigdyebuffer, 5*sequencingbuffer),Sodium acetate, polyethylene glycol, ethanol, 0.125MEDTA etc.
(I) from DNA sample, carry out the specific amplification of object fragment, reaction system is:
Reaction condition is:
(II) the single band qualification of PCR product, identifies object fragment after the method purifying of alcohol/EDTA again, and concrete grammar is:
(A) in every pipe 25 μ l systems, add 10 μ l sodium acetates (3M), 40 μ l polyethylene glycol (PDE8000), more than standing 1h.
(B) the centrifugal 40min of 3000g/min, 300r/min is inverted centrifugal 1min.
(C) every pipe adds 75% ethanol 70 μ l, the centrifugal 30min of 3000g/min, and 300r/min is inverted centrifugal 1min.
(D) after ethanol thoroughly volatilizees, every pipe adds 10 μ lddH2O dissolves.
(III) Bigdye sequencing reaction (ABI, BigDyeTeminatorV3.1), reaction system is:
Reaction condition is:
(IV) bigdye product purification, concrete grammar is:
(A) every pipe 10 μ l systems add 75% isopropyl alcohol 100 μ l, and more than standing 30min, the centrifugal 30min of 3000g/min, 300r/min fallsPut centrifugal 1min.
(B) add 75% ethanol (containing 0.125MEDTA, mixing with 75% ethanol with the volume ratio of 70:2), the 70 μ l containing EDTA, 3000g/minCentrifugal 15min, the centrifugal 1min of 300r/min.
(C) after ethanol thoroughly dries, add 10 μ lHIDI, after thermal denaturation (95 DEG C of 5min, ice bath 5min), turn order-checking plate, upper surveyOrder instrument.
Result is understood: peak figure is by Chromas software sectional drawing in order-checking, wherein,
Fig. 1-top represents that ADRB1rs1801253 site offside complementary strand allelotype is G, and patient's genotype is CC type, figure1-middle part represents that the genotype in ADRB1rs1801253 site is heterozygous CG type, and Fig. 1-bottom represents ADRB1rs1801253 site pairPlan complementary strand allele is C, and patient's genotype is GG type.
Fig. 2-top represents that GRK5rs17098707 loci gene type is AA type, and Fig. 2-bottom represents the base in GRK5rs17098707 siteBecause type is heterozygous AT type.
Improvement sequence measurement testing result statistics is as shown in table 1:
Table 1. is improved sequence measurement result statistics
Through traditional sequence measurement qualification, accuracy is 100%.
In embodiment 1, the nucleotide sequence of the primer is as shown in table 2:
Table 2. is improved the primer sequence for increase object fragment and order-checking relating in sequence measurement
The detection method of the order-checking of the improvement for ADRB1rs1801253 and GRK5rs17098707 site provided by the invention checks order in traditionOn the basis of method, optimize design of primers, pcr amplification primer can be used as sequencing primer simultaneously, has reduced design of primers cost, has simplified operation.Meanwhile, method optimizing reaction system on the basis of traditional sequence measurement of improvement order-checking provided by the invention, before the correct accuracy of guaranteePut the consumption that has greatly reduced sequencing reagent, in Bigdye sequencing reaction system, using 2.5 × Bigdyebuffer is only 0.25 μ l, is mark1/16 of mutatis mutandis amount 4 μ l, greatly reduce testing cost. In addition, the method for improvement order-checking provided by the invention is at the base of traditional sequence measurementOptimum Operation flow process on plinth, from sample process to obtaining testing result, has simplified operating procedure in 1 day greatly, has realized fast detecting, symbolClose the needs that clinical reagent uses.
The method of embodiment 2Taqman probe Genotyping determine patients with heart failure ADRB1rs1801253 and GRK5rs17098707 manyThe genotype in state property site
50 routine patients with heart failure ADRB1rs1801253 and GRK5rs17098707 more than determining by the method for Taqman probe GenotypingThe genotype of pleomorphism site, concrete scheme is:
The sequence of all primers and probe is synthesized by Jikang Biotechnology Co Ltd, Shanghai, after primer and probe sequence are synthetic, uses Tris-EDTASolution is dissolved as the concentration of 100uM/L and preserves. Other main agents is 2*GenotypeMasterMix (buying the company in American AB I).
The preparation (1/2384 orifice plate consumption) of Genotyping reaction reagent:
Above reagent mix is even, is sub-packed in 384 orifice plates with 4.5 μ l, and every hole adds experimenter's DNA profiling 0.6 μ l, covers hyaline membrane,Sealing. After centrifugal one minute of 3000g/min, upper machine carry out PCR reaction (AppliedBiosystems7900 real-time PCR,ABI, the U.S.)
Setting parameter is:
Later 384 orifice plates of amplification are put into AppliedBiosystems7900HT and are carried out Genotyping program.
Result is understood: somatotype result is understood (Fig. 3-4) with SDS2.3 software. As shown in the figure: the loose point in bottom right represents Fam only to be detected in sampleFluorescence, experimenter's genotype is wild type (bottom right). In the loose some representative sample of upper left, Vic fluorescence only detected, experimenter's genotype is for isozygotyingMutator type (upper left). In middle loose some representative sample, Fam fluorescence both detected, and Vic fluorescence detected again, experimenter's genotype isWild sudden change mixes genotype (centre).
After generation sequence verification, show: result and the sequencing and typing result of Taqman probe somatotype are in full accord.
In embodiment 2, the nucleotide sequence of the primer and probe is as shown in table 3:
The primer sequence and the probe sequence that in table 3.Taqman probe classifying method, relate to
The technology of the Taqman probe Genotyping that invention provides, the method for Taqman probe somatotype provided by the invention, at primer and probeOn sequences Design and experimental program, make optimization, ensureing under the prerequisite of experimental result accuracy, standard consumption 10ul reaction system/people'sDetect the reaction system that consumption reduces to 5ul/ people, make experiment reagent be reduced to 1/2 of standard consumption,, effectively save testing cost, energyBetter meet the demand of clinical practice. Meanwhile, as preferably 3 ' the non-fluorescent quenching group of end is MGB modification group, ensures to detectSpecificity and success rate, ensured the requirement of clinical practice to accuracy.
Claims (10)
1. one kind is detected ADRB1-Arg389Gly (rs1801253) and/or GRK5-Gln41Leu (rs17098707) pleomorphism siteThe reagent of Genotyping.
2. reagent as claimed in claim 1, is characterized in that, described gene polymorphism sites is (1) ADRB1-Arg389Gly, or (2) GRK5-Gln41Leu (rs17098707) (rs1801253).
3. the reagent as described in as arbitrary in claim 1 or 2, is characterized in that, described reagent comprises: 1, for comprising above-mentioned 2 SNP sitesThe specificity amplification primer pair of corresponding target sequence; And/or 2, identification pleomorphism site detector probe.
4. reagent as claimed in claim 3, is characterized in that, described specificity amplification primer is to being the oligonucleotide sequence of 14-30 nucleic acid,The object fragment that energy specific amplification comprises above-mentioned pleomorphism site.
5. the reagent as described in as arbitrary in claim 3 or 4, is characterized in that, described detector probe is the oligonucleotide sequence of 14-50 nucleic acid,Can specific and above-mentioned pleomorphism site specific hybrid.
6. the arbitrary described reagent of claim 1-5, in the purposes of preparing in kit, is characterized in that, described kit is suffered from for assessment of heart failurePerson's prognosis and/or beta-blocker medicine treatment curative effect.
7. for assessment of a kit for patients with heart failure prognosis and/or beta-blocker medicine treatment curative effect, it is characterized in that described examinationAgent box comprises the arbitrary described reagent of claim 1-5.
8. one kind is detected ADRB1-Arg389Gly (rs1801253) and/or GRK5-Gln41Leu (rs17098707) pleomorphism siteThe method of Genotyping, it is characterized in that, described method comprises utilizes the arbitrary described reagent of claim 1-5 to examine sampleSurvey.
9. method as claimed in claim 8, is characterized in that, comprises (1) improvement sequence measurement; (2) Taqman probe GenotypingMethod (3) other based on specific amplification and/or identify above pleomorphism site and the method for sequence around, as high-resolutionThe method of solubility curve, liquid phase DNA chip classifying method, solid phase DNA chip classifying method, for the PCR that comprises object fragmentThe analytical technique of mass spectrum of product, restriction enzyme typing method, allele hybridization technique, oligonucleotides connect experiment etc.
10. the method as described in as arbitrary in claim 8-9, is characterized in that, described sample is selected from: PBC, leucocyte, serum,Urine sample, saliva, body fluid and/or (biopsy) tissue samples. Preferably, described sample carries out purifying in advance, for example, separate total DNA.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610029394.2A CN105586406A (en) | 2016-01-15 | 2016-01-15 | Method for detecting gene polymorphism of ADRB1 and GRK5 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610029394.2A CN105586406A (en) | 2016-01-15 | 2016-01-15 | Method for detecting gene polymorphism of ADRB1 and GRK5 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN105586406A true CN105586406A (en) | 2016-05-18 |
Family
ID=55926306
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610029394.2A Pending CN105586406A (en) | 2016-01-15 | 2016-01-15 | Method for detecting gene polymorphism of ADRB1 and GRK5 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN105586406A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| 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 |
| CN106520979A (en) * | 2016-11-30 | 2017-03-22 | 武汉海吉力生物科技有限公司 | Nucleic acid, kit and method for detecting G1165C polymorphism of human ADRB1 gene |
| CN110607360A (en) * | 2019-09-19 | 2019-12-24 | 华中科技大学同济医学院附属同济医院 | Primer group and kit for assessing chronic heart failure prognosis and method for assessing chronic heart failure prognosis |
| CN113421648A (en) * | 2021-06-21 | 2021-09-21 | 中国科学院北京基因组研究所(国家生物信息中心) | Model for predicting risk of ejection fraction retention type heart failure |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102021238A (en) * | 2010-06-08 | 2011-04-20 | 广州益善生物技术有限公司 | ADRB1 (adrenergic, beta-1) gene SNP (Single Nucleotide Polymorphism) detection specificity primer and liquid phase chip |
| CN102876785A (en) * | 2012-09-13 | 2013-01-16 | 周宏灏 | Kit for detecting beta 1 receptor gene polymorphism by pyro-sequencing method and method |
| CN103525899A (en) * | 2012-07-02 | 2014-01-22 | 中国科学院上海生命科学研究院 | Type 2 diabetes susceptibility genetic locus as well as detection method and kit thereof |
-
2016
- 2016-01-15 CN CN201610029394.2A patent/CN105586406A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102021238A (en) * | 2010-06-08 | 2011-04-20 | 广州益善生物技术有限公司 | ADRB1 (adrenergic, beta-1) gene SNP (Single Nucleotide Polymorphism) detection specificity primer and liquid phase chip |
| CN103525899A (en) * | 2012-07-02 | 2014-01-22 | 中国科学院上海生命科学研究院 | Type 2 diabetes susceptibility genetic locus as well as detection method and kit thereof |
| CN102876785A (en) * | 2012-09-13 | 2013-01-16 | 周宏灏 | Kit for detecting beta 1 receptor gene polymorphism by pyro-sequencing method and method |
Non-Patent Citations (1)
| Title |
|---|
| LIGGETT SB ET AL.: "A GRK5 polymorphism that inhibits beta-adrenergic receptor signaling is protective in heart failure", 《NAT MED》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| 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 |
| CN106520979A (en) * | 2016-11-30 | 2017-03-22 | 武汉海吉力生物科技有限公司 | Nucleic acid, kit and method for detecting G1165C polymorphism of human ADRB1 gene |
| CN110607360A (en) * | 2019-09-19 | 2019-12-24 | 华中科技大学同济医学院附属同济医院 | Primer group and kit for assessing chronic heart failure prognosis and method for assessing chronic heart failure prognosis |
| CN113421648A (en) * | 2021-06-21 | 2021-09-21 | 中国科学院北京基因组研究所(国家生物信息中心) | Model for predicting risk of ejection fraction retention type heart failure |
| CN113421648B (en) * | 2021-06-21 | 2024-04-23 | 中国科学院北京基因组研究所(国家生物信息中心) | Model for predicting ejection fraction retention type heart failure risk |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN105586406A (en) | Method for detecting gene polymorphism of ADRB1 and GRK5 | |
| KR20030081339A (en) | Method of determining a chemotherapeutic regimen based on ercc1 and ts expression | |
| CN104328164A (en) | Kit for detecting human EGFR gene mutation by using fluorescence probe hybridization method | |
| CN109082464B (en) | Primer group and kit for detecting hypertension drug metabolism related genes | |
| CN104232781A (en) | TaqMan probe real-time fluorescence PCR (Polymerase Chain Reaction) method for detecting HLA (Human Leukocyte Antigen)-B*5801 alleles | |
| CN111118138A (en) | Kit and method for detecting polymorphism of folate metabolism ability genes MTHFR and MTRR | |
| CN101363054B (en) | Method for detecting hereditary hearing loss relative connexin 26 gene GJB2 mutation and kit for detection | |
| CN104293932A (en) | Method for detecting HLA-B * 5801 allele based on real-time fluorescence PCR | |
| JP2009511026A5 (en) | ||
| CN109234370B (en) | MUT gene mutation detection kit | |
| CN109929927A (en) | A kind of accurate medication gene tester of depression, primer combination of probe and kit | |
| CN106755343A (en) | Cancer of pancreas Prognosis molecular marked compound | |
| CN105331623A (en) | MTB (mycobacterium tuberculosis) rpoB mutant gene and application thereof | |
| CN108130362A (en) | Kit and application for EGFR genetic mutation detection | |
| CN109182528B (en) | Glioblastoma multiforme auxiliary diagnosis and prognosis evaluation kit based on ITGB5 gene and use method thereof | |
| WO2009129693A1 (en) | Probes, liquidchip-based products, and methods for detection of egfr gene mutation | |
| CN106755309A (en) | Application of the molecular marked compound in cancer of pancreas prognosis evaluation product is prepared | |
| Lucht et al. | Influence of DRD2 and ANKK1 genotypes on apomorphine-induced growth hormone (GH) response in alcohol-dependent patients | |
| CN104404044B (en) | Detection method and its application with the susceptible relevant ANRIL gene extrons sub-district mononucleotide polymorphic site of myocardial infarction | |
| JPWO2006137487A1 (en) | Gene polymorphism useful for predicting reactivity to β-blockers | |
| CN106868197B (en) | Application of the gene marker in Huppert's disease | |
| CN111154842A (en) | Composition, kit and method for detecting polymorphism of aspirin resistance gene of human | |
| CN105586405A (en) | Method for detecting gene polymorphism of ADRB2 and GRK5 and clinical application | |
| CN112695083B (en) | Nucleic acid composition and kit for detecting gene polymorphism of medicine for hypertension | |
| CN111378739B (en) | Detection site for detecting CYP2C19 gene polymorphism, primer composition and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20160518 |