CN106947829A - KCNQ1OT1 genes, the application of aHIF genes and its primer and the kit for diagnosis of coronary heart disease - Google Patents
KCNQ1OT1 genes, the application of aHIF genes and its primer and the kit for diagnosis of coronary heart disease Download PDFInfo
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Abstract
Kit the invention provides the application of a kind of KCNQ1OT1 genes, aHIF genes and its primer and for diagnosis of coronary heart disease, is related to the technical field of gene diagnosis.The KCNQ1OT1 genes and aHIF genes that are there is provided using the present invention carry out the diagnosis of coronary heart disease, the characteristics of with sensitivity height, high specificity, short diagnostic window phase, diagnosis can be made in invasioning delitescence or early stage, and fast and effectively can quantitatively be detected, diagnosis is can be not only used for, the state of an illness and curative effect monitoring can be carried out again.
Description
Technical field
The present invention relates to gene diagnosis technical field, more particularly, to a kind of KCNQ1OT1 genes, aHIF genes and its draw
The application of thing and the kit for diagnosis of coronary heart disease.
Background technology
At present, angiocardiopathy (hypertension, coronary heart disease, congestive heart failure, apoplexy etc.) is still human health
Number one killer.According to statistics, 1,007 million peoples die from angiocardiopathy every year in worldwide.Although some existing preventions
And the measure of Primary treatment angiocardiopathy, but it is still Med Biol fundamentally to prevent and cure angiocardiopathy
Great difficult problem.Therefore, find the mark of angiocardiopathy and accurate detection is carried out to it turns into angiocardiopathy in early days
Diagnosis and the urgent task for the treatment of.
Substantial amounts of research shows that in disease of cardiovascular system evolution occurs for non-coding nucleic acid (non-coding RNA)
In play extremely important effect, be prevention and treatment potential target spot.Non-coding nucleic acid includes microRNA and lncRNA etc., non-
The research of code nucleic acid has very important significance for these major diseases.Non-coding nucleic acid participates in the hair of heart and blood vessel
Educate, the disease such as its unconventionality expression and atherosclerosis, coronary heart disease, heart failure, arrhythmia cordis, congenital heart disease and hypertension
Disease is closely related.Non-coding nucleic acid has specific express spectra in various angiocardiopathies.
Compared with conventional protide polysaccharide label diagnosis, carrying out medical diagnosis on disease with non-coding nucleic acid has sensitivity
Height, high specificity, diagnostic window phase are short (can make diagnosis in invasioning delitescence or early stage), and can carry out quantitative detection, both
Available for diagnosing, the remarkable advantages such as the state of an illness and curative effect evaluation can be carried out again.But, non-coding nucleic acid is in the diagnosis of heart disease
Application still have some deficits, it is still desirable to it is further to explore and study.
In view of this, it is special to propose the present invention.
The content of the invention
First purpose of the present invention is to provide the application of KCNQ1OT1 genes, and second object of the present invention is to carry
For the application of the primer of KCNQ1OT1 genes, third object of the present invention is to provide the application of aHIF genes, of the invention
4th purpose is the application for the primer for providing aHIF genes, and the 5th purpose of the invention is to provide to be used to diagnose coronary disease
The kit of disease, to alleviate the present situation that non-coding nucleic acid in the prior art studies deficiency in the diagnosis of coronary heart disease.
The invention provides application of the KCNQ1OT1 genes in the kit for diagnosis of coronary heart disease is prepared.
In addition, being prepared present invention also offers the primer for specific detection KCNQ1OT1 genes for diagnosing coronary disease
Application in the kit of disease.
Further, the primer for specific detection KCNQ1OT1 genes is：
KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)；
KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3).
In addition, present invention also offers a kind of kit for diagnosis of coronary heart disease, the kit includes KCNQ1OT1
The primer of gene, the primer of the KCNQ1OT1 genes is：
KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)；
KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3).
In addition, the application present invention also offers aHIF genes in the kit for diagnosis of coronary heart disease is prepared, described
AHIF genes have the sequence as shown in SEQ ID NO.4.
In addition, being prepared present invention also offers the primer for specific detection aHIF genes for diagnosis of coronary heart disease
Application in kit, the aHIF genes have the sequence as shown in SEQ ID NO.4.
Further, the primer for specific detection aHIF genes is：
aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；
aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
In addition, present invention also offers a kind of kit for diagnosis of coronary heart disease, the kit includes aHIF genes
Primer, the primer of the aHIF genes is：
aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；
aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
In addition, present invention also offers a kind of kit for diagnosis of coronary heart disease, the kit includes KCNQ1OT1
The primer of gene and the primer of aHIF genes,
The primer of the KCNQ1OT1 genes is：
KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)；
KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3)；
The primer of the aHIF genes is：
aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；
aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
Further, using the primer and the primer of the aHIF genes of the KCNQ1OT1 genes, the polymerase of progress
Chain reaction condition is：Template denaturation condition is 95 DEG C, 3 minutes, then circulates 95 DEG C, 5 seconds, 55 DEG C, 35 seconds, totally 40 by each
Circulation, extension of time is 50 seconds after last time is circulated, and taking-up is placed in 4 DEG C of preservations.
The diagnosis of coronary heart disease is carried out using the KCNQ1OT1 genes that provide of the present invention and aHIF genes, with sensitivity it is high,
The characteristics of high specificity, short diagnostic window phase, diagnosis can be made in invasioning delitescence or early stage, and can carry out fast and effectively
Quantitative detection, can be not only used for diagnosis, the state of an illness and curative effect monitoring can be carried out again.
Brief description of the drawings
, below will be to specific in order to illustrate more clearly of the specific embodiment of the invention or technical scheme of the prior art
The accompanying drawing to be used needed for embodiment or description of the prior art is briefly described, it should be apparent that, in describing below
Accompanying drawing is some embodiments of the present invention, for those of ordinary skill in the art, before creative work is not paid
Put, other accompanying drawings can also be obtained according to these accompanying drawings.
Fig. 1 is KCNQ1OT1 gene qPT-PCR quantitative analysis figures；
Fig. 2 is aHIF gene qPT-PCR quantitative analysis figures；
Fig. 3 is heart disease patients gene qPT-PCR quantitative analysis figure of the KCNQ1OT1 genes in non-CAD；
Fig. 4 is heart disease patients gene qPT-PCR quantitative analysis figure of the aHIF genes in non-CAD.
Embodiment
Technical scheme is clearly and completely described below in conjunction with embodiment, it is clear that described reality
It is a part of embodiment of the invention to apply example, rather than whole embodiments.Based on the embodiment in the present invention, the common skill in this area
The every other embodiment that art personnel are obtained under the premise of creative work is not made, belongs to the model that the present invention is protected
Enclose.
Although the researchs for being directed to coronary heart disease existing many at present, it has been found that some lncRNAs related to coronary heart disease,
Diagnostic lncRNA is needed with certain specificity and sensitivity, therefore, for the lncRNA for diagnosis of coronary heart disease
Correlative study, still have deficiency.
Inventor solves above-mentioned technical problem, it is proposed that the solution of the present invention.
KCNQ1OT1 genes of the present invention are NR_002728.3, KCNQ1OT1 genes in NCBI VERSION numberings
Length be 91671bp.In view of KCNQ1OT1 gene orders are long, in this application, the part of KCNQ1OT1 genes has been intercepted
Fragment is as representative, and the Partial Fragment of KCNQ1OT1 genes is as shown in SEQ ID NO.1.
Sequence shown in SEQ ID NO.1 as KCNQ1OT1 genes representative, it may have KCNQ1OT1 genes are in the application
In effect.That is, sequence also can be used in preparing the kit for diagnosis of coronary heart disease shown in SEQ ID NO.1；For special
Property sequence shown in detection SEQ ID NO.1 primer, also can be used in preparing the kit for diagnosis of coronary heart disease.
The primer of KCNQ1OT1 genes of the present invention, it should be understood that be capable of specific detection KCNQ1OT1 genes
All primers, not only including the primer of KCNQ1OT1 gene complete sequences can be detected, also including KCNQ1OT1 genes can be detected
The primer of Partial Fragment.Wherein, the primer sequence shown in SEQ ID NO.2 and SEQ ID NO.3 is that Detection results are preferable
The primer of KCNQ1OT1 genes, and it is not construed as the limitation of the primer to KCNQ1OT1 genes.
The primer of aHIF genes of the present invention, it should be understood that be capable of all drawing of specific detection aHIF genes
Thing, not only including the primer of aHIF gene complete sequences can be detected, also including the primer of aHIF Gene Partial fragments can be detected.
Wherein, the primer sequence shown in SEQ ID NO.5 and SEQ ID NO.6 is the primer of the preferable aHIF genes of Detection results, and
It is not construed as the limitation of the primer to aHIF genes.
Kit of the present invention, the primer except including KCNQ1OT1 genes, either the primer of aHIF genes or
Outside the primer of KCNQ1OT1 genes and the primer of aHIF genes, it can also include being used for other correlation examinations that qRT-PCR reacts
Agent；It is preferred that, it can also include being used for the related reagent that RNA is extracted.
CAD (coronary atherosclerotic heart disease, CAD) of the present invention is coronary heart disease.
Unless otherwise indicated, the operating method being related in following examples is conventional method, and the reagent being related to is conventional
Commercial reagent.
The screening of embodiment 1KCNQ1OT1 genes and aHIF genes
1. preliminary screening and the closely related lncRNA of coronary heart disease
Extensive consulting literatures and lncRNA Relational databases, therefrom preliminary screening go out and coronary atherosclerotic heart disease
Closely related lncRNA (at least filtering out 30 key lncRNA).
2. Preliminary Identification lncRNA and the correlation of coronary heart disease
Collect clinical sample and set up case exclusion standard.Select case：The CAD and control group for receiving coronary angiography suffer from
Person, the relevant clinical data of patient is complete.
According to the purpose lncRNA filtered out, real-time fluorescence quantitative PCR detection gene expression amount range specifications：Using real-time
Quantifying PCR method verifies that key lncRNA is notable in patient CAD and control group PMNC (PBMC) sample
Property differential expression and the method such as indicatrix (ROC) and TG-AUC (AUC) detect to assess lncRNA in blood plasma as CAD
New diagnostic instrument feasibility.
3. verify accuracys and specific diagnosis of the crucial lncRNA in coronary heart disease
Collect CAD and non-CAD heart disease patients PMNC (PBMC) sample：Extract outside patient to be detected
All blood 5ml are placed in EDTA- anticoagulant tubes, and human peripheral blood single nucleus cell is separated using lymphocyte separation medium.
Sample cDNA templates are obtained：Human PBMC's cell of separation is extracted into RNA, reverse transcription synthesis cDNA in TriZol methods
Template.
Control group and gene expression difference in CAD groups PBMC, and CAD and non-CAD are detected by real-time fluorescence quantitative PCR
Gene expression dose compares in heart disease patients PBMC, thus verify key lncRNA as diagnosis CAD accuracy and
Specificity.
Therefrom filtered out 2 key lncRNA can specific diagnosis go out coronary heart disease, respectively KCNQ1OT1 and aHIF.
The primer of embodiment 2KCNQ1OT1 genes and aHIF genes is determined
Must have as the primed probe of diagnosis combine and expression specificity, stable existence, experiment are simple etc.
Feature, consistent results can be obtained through experiment is repeated several times.RT-PCR is carried out to normal and pathology sample with primed probe, contrasted
It expresses situation of change (whether can significantly judge its differential expression) and its repeated (whether process of the test is stablized), so as to sentence
Breaking, whether it can fast and accurately be diagnosed.To sum up, the present embodiment is from the standard of probe：
(1) significant differential expression can be detected in corresponding control group sample, is easy to clear analysis.
(2) differential expression between sample repeats that stable numerical value is presented through many experiments, occurs without and is widely varied, no
To then the accuracy of diagnosis be influenceed.
(3) probe in itself specific good, is not present mispairing in autogene group.
(4) there is general annealing temperature (55-60 degree), simplify experimentation, be easy to batch experiment.
Based on above principle, the present embodiment determines the primer of KCNQ1OT1 genes, and particular sequence is：
KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)
KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3)
And the primer of aHIF genes, particular sequence is：
aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；
aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
The kit of the diagnosis of coronary heart disease of embodiment 3
The kit that the present embodiment is provided includes the primer of KCNQ1OT1 genes, and particular sequence is：
KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)
KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3)
The kit of the diagnosis of coronary heart disease of embodiment 4
The kit that the present embodiment is provided includes the primer of aHIF genes, and particular sequence is：
aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；
aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
The kit of the diagnosis of coronary heart disease of embodiment 5
The kit that the present embodiment is provided includes：The primer of KCNQ1OT1 genes, and aHIF genes primer.
Wherein, the primer of KCNQ1OT1 genes is：
KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)
KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3)
The primer of aHIF genes is：
aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；
aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
The application method of the kit of the diagnosis of coronary heart disease of embodiment 6
The application method of the kit including embodiment 2,3 and 4 is present embodiments provided, is comprised the following steps：
Step (a), the extraction of patient's PMNC template DNA to be detected：Extract patient's peripheral blood to be detected
5ml is placed in EDTA- anticoagulant tubes, and human peripheral blood single nucleus cell is separated using lymphocyte separation medium.TriZol methods are extracted
RNA, reverse transcription synthesis cDNA templates.
Step (b), polymerase chain reaction：
Reaction system：
Reaction condition：Template denaturation condition is 95 DEG C, 3 minutes, 5 seconds, 55 DEG C, 35 seconds, common then by each 95 DEG C of circulation
40 circulations, extension of time is 50 seconds after last time is circulated, and taking-up is placed in 4 DEG C of preservations.
In addition, present invention also offers the experiment for applying above example, more fully to illustrate the effect of the present invention,
It is specifically described as follows.
The kit that Application Example 5 is provided, and the method that embodiment 6 is provided, are examined to control group and CAD patient
Survey.Testing result is as depicted in figs. 1 and 2.
Fig. 1 is KCNQ1OT1 gene qPT-PCR quantitative analysis figures, wherein, control group is that Normal group peripheral blood is single
Relative KCNQ1OT1 gene contents in nucleus；CAD contains for relative KCNQ1OT1 genes in CAD peripheral blood mononuclear cells
Amount.As seen from Figure 1, the content of KCNQ1OT1 genes is substantially less than control group in CAD peripheral blood in patients, therefore, it can make
To judge whether patient suffers from the foundation of coronary heart disease.
Fig. 2 is aHIF gene qPT-PCR quantitative analysis figures, wherein, control group is that the single core of Normal group peripheral blood is thin
Relative aHIF gene contents in born of the same parents；CAD is relative aHIF gene contents in CAD peripheral blood mononuclear cells.Can be with by Fig. 2
Find out, the content of aHIF genes is significantly higher than control group in CAD peripheral blood in patients, therefore, it can as judging whether patient suffers from
There is the foundation of coronary heart disease.
In addition, the kit that also Application Example 5 is provided, and the method that embodiment 6 is provided, non-CAD patient is carried out
Detection.Testing result is as shown in Figure 3 and Figure 4.
Fig. 3 is heart disease patients gene qPT-PCR quantitative analysis figure of the KCNQ1OT1 genes in non-CAD, wherein, control
Group is relative KCNQ1OT1 gene contents in Normal group PMNC；CAD is the single core of CAD peripheral blood in patients
Relative KCNQ1OT1 gene contents in cell；Valvular heart disease group is valvular heart disease (non-CAD other class heart diseases) peripheral blood in patients
Relative KCNQ1OT1 gene contents in mononuclearcell.As seen from Figure 3, KCNQ1OT1 genes in CAD peripheral blood in patients
Content is substantially less than control group, but the content of KCNQ1OT1 genes is compared no statistics with control group in Patients With Valvular Heart Disease peripheral blood
Difference is learned, therefore, KCNQ1OT1 genes can be used as the foundation for judging specific detection coronary heart disease.
Fig. 4 is heart disease patients gene qPT-PCR quantitative analysis figure of the aHIF genes in non-CAD, wherein, control group is
Relative aHIF gene contents in Normal group PMNC；CAD is phase in CAD peripheral blood mononuclear cells
To KCNQ1OT1 gene contents；Valvular heart disease group is that valvular heart disease (non-CAD other class heart diseases) single core of peripheral blood in patients is thin
Relative aHIF gene contents in born of the same parents.As seen from Figure 4, the content of aHIF genes is significantly higher than control in CAD peripheral blood in patients
Group, but the content of aHIF genes is compared no difference of science of statistics with control group in Patients With Valvular Heart Disease peripheral blood, therefore, aHIF genes
The foundation for judging specific detection coronary heart disease can be used as.
Moreover, the testing result of KCNQ1OT1 genes and aHIF genes be combined with each other, the accurate of detection can further improve
Property.
The diagnosis of coronary heart disease is carried out using the KCNQ1OT1 genes that provide of the present invention and aHIF genes, with sensitivity it is high,
The characteristics of high specificity, short diagnostic window phase, diagnosis can be made in invasioning delitescence or early stage, and can carry out fast and effectively
Quantitative detection, can be not only used for diagnosis, the state of an illness and curative effect monitoring can be carried out again.The diagnostic kit that the present invention is provided, will fill up state
The industry blank of interior diagnosis of coronary heart disease.
Finally it should be noted that：Various embodiments above is merely illustrative of the technical solution of the present invention, rather than its limitations；To the greatest extent
The present invention is described in detail with reference to foregoing embodiments for pipe, it will be understood by those within the art that：Its according to
The technical scheme described in foregoing embodiments can so be modified, or which part or all technical characteristic are entered
Row equivalent；And these modifications or replacement, the essence of appropriate technical solution is departed from various embodiments of the present invention technology
The scope of scheme.
SEQUENCE LISTING
<110>University Of Qingdao
<120>KCNQ1OT1 genes, the application of aHIF genes and its primer and the kit for diagnosis of coronary heart disease
<130> 2017
<160> 6
<170> PatentIn version 3.5
<210> 1
<211> 5000
<212> DNA
<213>People（Homo sapiens）
<400> 1
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agtaaaatac aggacagagt tataaagtgc ctgaactttg aatgtcccaa cccacacata 180
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ctcatacgga gtgcacaacc tagatccctc acatgctcag ttcacaatag gatttgctct 1080
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gtaaaatgat ggaaaaagat gacatgcaaa cagtaataag aaagctagag tgactgtact 1800
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aaactctgtc tctactaaaa attagctggg tatggtggca aatgcctgta gtcccagcta 1980
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gatgataaat gccaactgaa agtaaaagga tggaaaaaga tatatcatgt aagcagcaac 3480
cataagtaac ctgaagtgct ataataaagt caaaggagat tttaaattaa gaatgttact 3540
gaagttaaat atgaatatct cataatgata aaatgtcaat tcatgagaag atacaatgat 3600
tataaacata tattcaccta acaatagaga actaaaacat aagaagcaaa aactgacaga 3660
actgaacaga gaaatagcca attcaacaat agtagttgga gacttcaaca ctccactttc 3720
aataatgtgt agtacaagaa agaagatcaa taaggaaaca gaagatatga gcaacaccat 3780
aaaccaacta gactcaaaag gtaattatag agcacttcgc ccaacaacca acattgaata 3840
tacagtattt tcaagtgcac atgagacatt ctccaagaag gaccatatgc taagtcacga 3900
aacaaacctg cataaattta aaaggattga aataatacaa ggtatctttt tctgaatata 3960
atgaaataaa agtagaactc agtagcaaag aaattaggga aacatacaca tgtggaaatt 4020
aaacaacact acattcttaa ataatcaatg ggtgaaagaa ttcacaaggg aaatggaacg 4080
tactttgaga ttaatgaaaa tgaagataca acataacaga acttatggga tacatttaaa 4140
gcagtgtaaa gatggaattt tatagctatg aatgcctata tcaaaaaata aggcttcaaa 4200
tcagtaagcc aactctccac ctcaagaaac tagaaaaaga agagcaaact aaacctaaag 4260
caagtagaaa aaacaaaata ataaatatta gagtagaaat aaaatataga atataaaaat 4320
aataaagtca acaaatccaa aatttggttc tttgaaatta acaaaccttt gatagattgg 4380
taagagagag ggagagggag aagaaaagaa ggggaggaag aaagaggggg aggaggaaag 4440
acagagaggg aagtgcatga ggcaaagggg gagagagaaa actcaaatta ctaaaatcag 4500
aaatgaatga cagttttaca aattaaagca aaaaaacgag gctagctgca gtggctcaca 4560
cctgtaatcc cagcactttg ggaggccaag gcaggagaat tgcttgtggc caggagttcg 4620
agaattgctt gtggccagga gttcaagaat agcatgcaca acaaagcaag accctatctc 4680
tacaaaatat tttttgaata acagctggtg ttgtggtgca cacctgtcat aggctaaggg 4740
gcaaggatca cttgagacta ggagatcgag gttacagtga gctatgatta caccactcta 4800
ttccagcctg ggcaacagag agagactctg tttctcaaaa agaaaaggaa aaaggaatga 4860
atgagggaag tatactacca acctgacaga aataaaaagg attatgaagg aacactatga 4920
acaattttgt gccaataaat tataaaactt agatgaaatt gacaaattcc tagaaagatg 4980
aaaactatga aaactgacta 5000
<210> 2
<211> 19
<212> DNA
<213>Artificial sequence
<400> 2
atagtagttg gagacttca 19
<210> 3
<211> 20
<212> DNA
<213>Artificial sequence
<400> 3
actgtatatt caatgttggt 20
<210> 4
<211> 2051
<212> DNA
<213>People（Homo sapiens）
<400> 4
tgtcctccat tgtaagataa aaagagctac ctaagagatc tgtggctcag ttccttttgt 60
tcagtatata tacctcaggg taaaggacct aaggctctgg cacttcctac ataatttgtt 120
cataaaaata attttctgta cctgtaggtg cagggattca tagtgatttt tctgttttaa 180
aaaatatctt tctacactgt ctgattttta aaaatgagat tatcagtata aaagaaatac 240
agctattttg taaaagaaaa atacaggaga aaaaggataa agctactttg ttttaaaacg 300
cagtatcatt aaaacaaaaa caaaccaaat ttgttctaag tttgacttta gagtcaggaa 360
acttaagctt acatttttgg tctgccatct attactttta aagcttgggc aaattattca 420
tttgaagtct aaatttatta atctgttaat gggaacagat tagaaatctt cagagaagct 480
ctagcctttg taacattgtg actataatgc tgagaactgc ttcactcatc ccattcatat 540
tttaaaaata ctaatatttt gtgtttgagc attttaatag gctcagaaac ttaaaaatga 600
tgtttctttt ctaacaacat actcttttca atgggatatt atggttgtta ttatttaaca 660
tgacatttag ggactcaaca tacattaagg tgatggcact aagataaatg tagaaataac 720
cagtaccatg taattttcat aagtgcttaa attgttggta aacaatttta tgagttggag 780
gtgttgaagc aaatattatt aatatttgaa cataaaagct gatcaaaggg gcctggtcca 840
cagaagatgt ttatttgatg taacaaaaca atacagttag tgttagatcc aaccacaaag 900
agcaaaagga atgaaaaatt tgtacaatgt acatagaaaa aacaagatat ttactgtgac 960
aactatatat tcctaaaata atgcttctaa aattactcaa ttattgaaat ctacatgaaa 1020
aaaaggatgt taatagcgac aaagtgcata aaatcaaaca ttgtattttg agcaaattaa 1080
catactaggc aattttgcta agaatgcatg attttttttt tcttgtttac agtctgctca 1140
aaatatcttt ataccaacag ggtaggcaga acatttaggt ttaatatcag ttacacaata 1200
ttagcataaa cttccacaac tacatagggt attgttttct tttgagctgg caaagtgact 1260
atagaaacat cagatgattt ctctgaattg agaattttat ccaaataaat gccacatacc 1320
ttctagatat atgcatatct ttctatatta tgtaaatggc tttacccatt taaataataa 1380
accatacagc atttaagaat cattattata tgattaacaa tgtcatgttc caggtttaac 1440
aatttcatag gccaaaaaaa atttcttctt aaaaactagt tttataaacg cagaatatat 1500
tccatgagta actgctggta ttttttaaga aaatatattg tgcaattgtg gctaccacgt 1560
actgctggca aagcattatt atttatgtaa aatgtgaaaa aaaaggtgta aaaatttttc 1620
aactgcctat gatcatgatg aaaggttact gccttcttac aaaaattata ttggcatctt 1680
cttaaaaata attcgaaaaa gggataaact ccctagccaa aaataaataa ataaaaaggt 1740
gcatttttta aaatgatgct actgcaatgc aatggtttaa ataccaaaaa actgagaaaa 1800
tgagctgtct gtgatccagc attaaagaac atactaaaaa agagcattaa tgtaaattaa 1860
gtagaaaggg gatcaaaatt gaactaacca agtttgtgca gtattgtagc caggcttcta 1920
aaattagatg tagaaaatat aaatagactg ctttaggtaa tgagccacca gtgtccaaaa 1980
aaaggaatga aattaagaaa aagctcagtt aacttgatcc aaagctctga gtaattcttc 2040
accctgcagt a 2051
<210> 5
<211> 19
<212> DNA
<213>Artificial sequence
<400> 5
ggtctgccat ctattactt 19
<210> 6
<211> 20
<212> DNA
<213>Artificial sequence
<400> 6
tctcagcatt atagtcacaa 20
Claims (10)
- Application of the 1.KCNQ1OT1 genes in the kit for diagnosis of coronary heart disease is prepared.
- 2. application of the primer for specific detection KCNQ1OT1 genes in the kit for diagnosis of coronary heart disease is prepared.
- 3. application according to claim 2, it is characterised in that the primer for specific detection KCNQ1OT1 genes For：KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)；KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3).
- 4. a kind of kit for diagnosis of coronary heart disease, it is characterised in that the kit includes the primer of KCNQ1OT1 genes, The primer of the KCNQ1OT1 genes is：KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)；KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3).
- Application of the 5.aHIF genes in the kit for diagnosis of coronary heart disease is prepared, it is characterised in that the aHIF genes tool Just like the sequence shown in SEQ ID NO.4.
- 6. application of the primer for specific detection aHIF genes in the kit for diagnosis of coronary heart disease is prepared, its feature It is that the aHIF genes have the sequence as shown in SEQ ID NO.4.
- 7. application according to claim 6, it is characterised in that the primer for specific detection aHIF genes is：aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
- 8. a kind of kit for diagnosis of coronary heart disease, it is characterised in that the kit includes the primer of aHIF genes, described The primer of aHIF genes is：aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
- 9. a kind of kit for diagnosis of coronary heart disease, it is characterised in that the kit includes the primer of KCNQ1OT1 genes With the primer of aHIF genes,The primer of the KCNQ1OT1 genes is：KCNQ1OT1-F：5’-ATAGTAGTTGGAGACTTCA-3’(SEQ ID NO.2)；KCNQ1OT1-R：5’-ACTGTATATTCAATGTTGGT-3’(SEQ ID NO.3)；The primer of the aHIF genes is：aHIF-F：5’-GGTCTGCCATCTATTACTT-3’(SEQ ID NO.5)；aHIF-R：5’-TCTCAGCATTATAGTCACAA-3’(SEQ ID NO.6).
- 10. kit according to claim 9, it is characterised in that the primer of the application KCNQ1OT1 genes and described The primer of aHIF genes, the polymerase chain reaction condition of progress is：Template denaturation condition is 95 DEG C, 3 minutes, is then followed by each 95 DEG C of ring, 5 seconds, 55 DEG C, 35 seconds, totally 40 circulations, extension of time is 50 seconds after last time is circulated, and taking-up is placed in 4 DEG C of guarantors Deposit.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108315423A (en) * | 2018-04-10 | 2018-07-24 | 复旦大学附属妇产科医院 | Purposes of the serum excretion body aHIF as ovarian cancer diagnosis and the marker of outcome |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108315423A (en) * | 2018-04-10 | 2018-07-24 | 复旦大学附属妇产科医院 | Purposes of the serum excretion body aHIF as ovarian cancer diagnosis and the marker of outcome |
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