CN103667301A - Gene related to coronary heart disease, and in-vitro detection reagent, preparation or kit and application thereof - Google Patents
Gene related to coronary heart disease, and in-vitro detection reagent, preparation or kit and application thereof Download PDFInfo
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Abstract
The invention belongs to the relevant technical field of genes related to coronary heart disease in bioengineering, and relates to a gene related to coronary heart disease, and an in-vitro detection reagent, preparation or kit and application thereof. The gene has a nucleotide sequence as shown in SEQ ID NO:1. The gene related to coronary heart disease, and the in-vitro detection reagent, preparation or kit and application thereof are introduced in detail in the invention. The in-vitro detection kit of the gene related to coronary heart disease can be used for performing in-vitro detection on the polymorphism of the gene related to coronary heart disease, and can be also used for detecting, preventing, diagnosing and treating coronary heart disease, so that the incidence rate of coronary heart disease is reduced, thereby being of great significance in prevention and treatment of coronary heart disease.
Description
Technical field
the invention belongs to coronary heart disease dependent genes correlative technology field in biotechnology, relate to reagent, preparation or test kit, the application of a kind of coronary heart disease dependent genes and vitro detection thereof.
Background technology
epidemiology and clinical study show, coronary heart disease (coronary artery disease) is developed country and most developing countries grownup morbidity and main causes of death.Coronary heart disease morbidity and mortality ratio are the trend increasing year by year and have caused global concern.Many developing countries, along with the west of affluence and the mode of life of material conditions, expecting twenties 21 century coronary heart disease may become the rank the first disease of name of the whole world.Since the result of study announcement of the Frmainghma sixties in last century, people are also greatly improved to the understanding of coronary heart disease, have realized that coronary heart disease is not a kind of single factor disease, but by multiple endogenous and the common Complex Diseases determining of extrinsic factor, except hypertension, diabetes, smoking, outside the environmental factorss such as metabolism syndrome, the effect of inherited genetic factors in coronary heart disease generation evolution also obtained being further familiar with and having obtained the inherited genetic factors in coronary heart disease develops of attention to a certain degree and account for 40% to 60% progressively.
collagen protein (collagen) is a kind of unique texture that has, and a kind of structural protein of extracellular matrix, are extensively present in mammal tissue, account for the more than 30% of gross protein.The kind of collagen protein is more, and the kind of finding at present has 25 kinds at least, wherein structure and function ratio more clearly type be I type, II type, III type, IV type, V type.Different collagen distribution is in vivo different, and NTx is mainly distributed in muscle tendon, bone, tooth and various soft tissue organs; II Collagen Type VI is mainly distributed in hyaline cartilage and elastic cartilage; III Collagen Type VI is distributed in various soft tissue internal organs, distributes roughly the same with NTx; Type Ⅳ collagen is distributed in each histoorgan basilar membrane basement membrane; V Collagen Type VI is distributed in pericellular.
type Ⅳ collagen was separated by people such as Kefalides in 1966 first from glomerular basement membrane, was the primary structure composition of basilar membrane, accounted for 50% left and right of basement membrane components.The structure of type Ⅳ collagen is more special, (α 1-α 6) six chains, consists of, and the gene of these six α chains of encoding is with paired being positioned on 3 different karyomit(e)s of mode head to head.These six α chains are with α 1. α 1. α 2, α 3. α 4. α 4, and the mode of α 5. α 5. α 6 forms heterotrimer, and each α chain all comprises the 7S structural domain of 3 structural domain: N ends; Middle triple helical collagen structure territory, sweet amino-X-Y sequence that repetition is contained in this region (XY is generally proline(Pro) and oxyproline or Methionin and hydroxylysine), this tumor-necrosis factor glycoproteins forms in spiral process and plays an important role at collagen; The non-collagen structure territory (NC1) of C-terminal, NC1 is rich in halfcystine and Methionin.The discoveries such as Rodriguez, the disease relevant with type Ⅳ collagen abnormal protein is mainly because the sudden change of glycine in this protein adhesive prodomain is relevant, possible pathogenesis has following: glycine is a seed amino acid of molecular weight minimum, unique amino acid that can enter type Ⅳ collagen triple-helix structure center, glycine mutation can affect the formation of triple-helix structure or the secretion of type Ⅳ collagen, thereby affect the stability of basement membrane structure, the sudden change of glycine changes the CB3 plot structure in triple helical district in addition, hiding integrin protein binding site exposes, make type Ⅳ collagen proteolytic degradation or sex change, even the antibody in these sites may angiogenesis inhibiting.
in vascular system, type Ⅳ collagen is distributed in around the vascular smooth muscle cell in arterial wall middle level, is the main component of vessel wall basilar membrane.Public institute is known, the integrity of vessel wall and stability are most important for cardiovascular systems, this integrity and stability are by vessel wall basilar membrane, endotheliocyte and around between smooth muscle cell etc., mutually coordinate to obtain, if this coordinative role is damaged for some reason, can affect vascular system, result causes hemorrhage, oedema, the serious consequence such as inflammation and tissue ischemia, and these are all the bases that development occurs coronary heart disease.In addition, type Ⅳ collagen also plays an important role to the migration of cell, propagation, differentiation.
type Ⅳ collagen protein gene α 1(COL4A1), gene is positioned at 13q34, mRNA total length 6549bp, and coding IV Collagen Type VI α 1 chain, comprises 52 exons.COL4A1 gene is the new gene of finding in the genome-wide association study (GWAS) of a relevant coronary heart disease in the recent period, thinks that between genetic polymorphism and coronary heart disease, existence is associated.In addition, the people such as Y Yamada study first and show, the polymorphism of COL4A1 gene is relevant with the generation of myocardial infarction.For COL4A1 gene pleiomorphism and coded protein, the effect in cardiovascular systems generation evolution has caused people's concern gradually in recent years.Clinically this protein likely by priority application in the prediction index of the ill risk of coronary heart disease.
Summary of the invention
the invention provides reagent, preparation or test kit, the application of a kind of coronary heart disease dependent genes and vitro detection thereof, overcome the deficiency of above-mentioned prior art, it can effectively solve prior art undesirable problem of effect when prevention and treatment coronary heart disease.
one of technical scheme of the present invention realizes by following measures: a kind of coronary heart disease dependent genes is COL4A1 gene, and the nucleotide sequence of this gene has the sequence shown in SEQ ID NO:1.
two of technical scheme of the present invention realizes by following measures: the reagent of a kind of vitro detection coronary heart disease dependent genes according to claim 1, this reagent is for detection of the polymorphism in COL4A1 gene rs605143 site, and this reagent comprises following primer:
upstream primer: 5'AAAGCCATTGCTACCTCA3'
downstream primer: 5'CTGCTCCTGGTGACTCTG3'
the further optimization and/or improvements to two of foregoing invention technical scheme below:
mentioned reagent can be the reagent combining with restriction fragment length polymorphism analysis for polymerase chain reaction or the reagent combining with direct sequencing for polymerase chain reaction.
three of technical scheme of the present invention realizes by following measures: a kind of preparation or test kit that has the reagent of vitro detection coronary heart disease dependent genes, this test kit comprises pcr amplification enzyme and corresponding damping fluid.
four of technical scheme of the present invention realizes by following measures: a kind of reagent of vitro detection coronary heart disease dependent genes is in the preparation for the preparation of vitro detection coronary heart disease dependent genes or the application in test kit.
the further optimization and/or improvements to four of foregoing invention technical scheme below:
in mentioned reagent box, can comprise pcr amplification enzyme and corresponding damping fluid.
the present invention is to the reagent of coronary heart disease dependent genes and vitro detection thereof, preparation or test kit and should be used as detailed introduction, the test kit of this vitro detection coronary heart disease dependent genes is except can the polymorphism for vitro detection coronary heart disease dependent genes, can also for detection of, prevention, diagnosis or treatment coronary heart disease, thereby reduce the incidence of coronary heart disease, prevention and treatment coronary heart disease are significant.
Accompanying drawing explanation
fig. 1 is the examination schematic flow sheet of coronary heart disease dependent genes sequence variations in the present invention.
fig. 2 is part order-checking collection of illustrative plates of the present invention.
fig. 3 is the restriction enzyme mapping of rs605143 pleomorphism site of the present invention.Wherein as swimming lane 1 and 2 shows that the genotype in individual rs605143 to be measured site is G/G homozygote; Swimming lane 3,4,6 and 7 show that the rs605143 site of individuality to be measured is A/G heterozygote; Swimming lane 5 shows that the rs605143 site of individuality to be measured is A/A homozygote.
Embodiment
in order more clearly to understand the present invention, referring now to the following example and accompanying drawing, further describe the present invention, embodiment does not only limit the present invention in any way for explanation.The experimental technique of unreceipted actual conditions in embodiment, conventionally according to normal condition, carry out: as < < molecular clonings that people showed such as Sambrook: laboratory manual > > (New fork:Cold Spring Harbor Laboratory Press, 1989) condition described in, or the condition of advising according to manufacturer.
embodiment 1, and a kind of coronary heart disease dependent genes is COL4A1 gene, and the nucleotide sequence of this gene has the sequence shown in SEQ ID NO:1.The testing sample of the gene that contains COL4A1 can obtain from the cell from trier, and the cell of Tathagata autoblood, urine, saliva, gastric juice, hair, examination of living tissue and necrotomy material, preferably carrys out autoblood.
embodiment 2, a kind of reagent of vitro detection coronary heart disease dependent genes, and this reagent is for detection of the polymorphism in COL4A1 gene rs605143 site, and this reagent comprises following primer:
upstream primer: 5'AAAGCCATTGCTACCTCA3';
downstream primer: 5'CTGCTCCTGGTGACTCTG3';
so-called " gene pleiomorphism " refers in crowd, the difference that the nucleotide sequence of each genes of individuals exists.Those of ordinary skills are known, and pleomorphism site of the present invention is single nucleotide polymorphism (SNP) site, and in genome sequence, mononucleotide changes; The difference of nucleotide acid sequence can be embodied on DNA level or on rna level, so coronary heart disease dependent genes of the present invention can be embodied in DNA level, and on rna level, preferred DNA level, more preferably genomic dna.
research about complex character disease at present has two kinds of strategies, the method for linkage analysis and case-control study.Because coronary heart disease is a kind of multi-factor disease, be by the coefficient result of h and E factor, thereby adopt comparatively difficulty of linkage analysis.From hereditary angle analysis, coronary heart disease/myocardial infarction belongs to complex inheritance proterties disease.In the factor working of this disease, except environmental factors, the gene number working is estimated to reach up to a hundred, exists again interaction between each gene, and gene exists and interacts again and between environment.Coronary heart disease/myocardial infarction there is certain familial aggregation, but be more some Sporadic cases.So the present invention adopts case control study.Described case control study is exactly certain allelic frequency of (case and contrast) comparison in two groups of crowds that adopt random choose.
the present invention's statistical study by large sample in the Wei Weier clansman group of Xinjiang (471 routine patients with coronary heart disease and 624 example contrasts), through a large amount of experiments, last take conclusive evidence and proved that the COL4A1 gene with rs605143 pleomorphism site of the present invention is coronary heart disease dependent genes, the danger that the allelic carrier of G suffers from coronary heart disease has increased by 1.4 times compared with A allelotrope carrier
embodiment 3, as above-described embodiment preferably, this reagent is the reagent combining with restriction fragment length polymorphism analysis for polymerase chain reaction or the reagent combining with direct sequencing for polymerase chain reaction.The reagent that detects rs605143 site in test kit is according to the difference of detection method and difference, for example, while adopting polymerase chain reaction to detect rs605143 pleomorphism site with the restriction fragment length polymorphism analysis method of combining, in test kit, can contain restriction enzyme and corresponding restriction enzyme mapping, for example DraI restriction enzyme and corresponding restriction enzyme mapping.
embodiment 4, and a kind of preparation or test kit that has the reagent of vitro detection coronary heart disease dependent genes is characterized in that this test kit comprises pcr amplification enzyme and corresponding damping fluid.The present invention detects the test kit of the coronary heart disease dependent genes of rs605143 loci polymorphism can be for the polymorphism of vitro detection coronary heart disease dependent genes; The test kit of the coronary heart disease dependent genes of rs605143 site mutation can for detection of, prevention, diagnosis or treatment coronary heart disease; The method of vitro detection coronary heart disease dependent genes can for detection of, prevention, diagnosis or treatment coronary heart disease.
embodiment 5, and a kind of reagent of vitro detection coronary heart disease dependent genes is in the preparation for the preparation of vitro detection coronary heart disease dependent genes or the application in test kit.The present invention detects the test kit of the coronary heart disease dependent genes of rs605143 loci polymorphism can be for the polymorphism of vitro detection coronary heart disease dependent genes; The test kit of the coronary heart disease dependent genes of rs605143 site mutation can for detection of, prevention, diagnosis or treatment coronary heart disease; The method of vitro detection coronary heart disease dependent genes can for detection of, prevention, diagnosis or treatment coronary heart disease.
embodiment 6, are with the difference of above-described embodiment, and test kit also comprises pcr amplification enzyme and corresponding damping fluid.
embodiment 7, the choosing of research object
cHD group (CAD group) 471 examples, are selected from January, 2006～2010 year March at No.1 Hospital Attached to Xinjiang Medical Univ.'s Heart center inpatient, meet the World Health Organization (WHO) about name and the Case definition of ischemic heart disease.Control group 624 examples, select in Xinjiang region crowd's chronic disease investigation in 2006 to 2011 without any heart disease person.Two groups are the crowd of the Uygur nationality, and age and sex composition, than all mating, have harmony.
inclusive criteria: CAD group: confirm to have coronary stricture>=50% at least and without otherwise heart trouble illness according to my institute percutaneous coronary radiography.Control group: select in Xinjiang region crowd's chronic disease investigation in 2006 to 2011 without any heart disease person. two groups of signature Informed Consent Forms before including research in.
rejecting standard: CAD group: the full person of clinical data and the one that simultaneously merges following disease are rejected.(as: dissection of aorta, rheumatic heart disease, Congenital Heart patient, MOFE and there is mental disorder can not cooperation person).Control group: 1, confirm to suffer from any heart disease person 2 in Xinjiang region crowd's chronic disease investigation in 2006 to 2010, have cardiovascular diseases family history person 3, drug addict 4, mentally disturbed, rejected.
whole coronary heart disease dependent genes sequence variations examination flow process is shown in Fig. 1, from case group, the routine patients with coronary heart disease of random choose 36 is as examination object, form 72 karyomit(e)s, this sample size can provide 95% power of a test the sequence variations that rare gene frequency is greater than 1% to be detected.All individualities are the Uygur nationality, and without any genetic connection.COL4A1 exon sequence according to announcing in NCBI, relatively obtains COL4A1 gene order by blast database.The method that detects polymorphism is to utilize PCR product direct Sequencing.Utilize Primer 5.0 software design PCR primers, amplification COL4A1 gene rs605143 pleomorphism site.Primer sequence is:
upstream primer: 5'AAAGCCATTGCTACCTCA3';
downstream primer: 5'CTGCTCCTGGTGACTCTG3'.
embodiment 8, the examination of coronary heart disease dependent genes sequence variations
pCR reaction system (50 μ l): genomic templates DNA 50ng is (from blood preparation, according to ordinary method by intraleukocytic DNA by the imitative method of phenol-atmosphere or with salting-out process, extract), 2*powder Taq PCR master mix 25ul, 21ul of deionized water, on, lower each 1ul of primer, in the reaction of the enterprising performing PCR of 96 hole PCR automatic circulation instrument, PCR loop parameter: 96 ℃ of denaturations 5 minutes; Through 94 ℃ 30 seconds, 60 ℃ 45 seconds, 72 ℃ 1 minute; After circulating 28 weeks, in 72 ℃, extend 10 minutes.PCR product carries out sequencing reaction after conventional purifying 2 times, usings PCR primer as sequencing primer.Order-checking completes on ABI 3500 sequenators.Fig. 2 display section order-checking collection of illustrative plates.Primer sequence is in Table 1.
result: take the Xinjiang, China Uygur nationality as research object, by a series of laboratory facilities, find that between COL4A1 gene rs605143 pleomorphism site and the crowd of Xinjiang, China Uygur nationality coronary heart disease, existence is associated.
embodiment 9, adopt PCR-RFLP method to detect the polymorphism in coronary heart disease dependent genes rs605143 of the present invention site
method: PCR reaction system (50 μ l): genomic templates DNA 50ng is (from blood preparation, according to ordinary method by intraleukocytic DNA by the imitative method of phenol-atmosphere or with salting-out process, extract), 2*powder Taq PCR master mix 25ul, 21ul of deionized water, on, lower each 1ul of primer, in the reaction of the enterprising performing PCR of 96 hole PCR automatic circulation instrument, PCR loop parameter: 96 ℃ of denaturations 5 minutes; Through 94 ℃ 30 seconds, 60 ℃ 45 seconds, 72 ℃ 1 minute; After circulating 28 weeks, in 72 ℃, extend 10 minutes.。Primer is as follows:
upstream primer: 5'AAAGCCATTGCTACCTCA3';
downstream primer: 5'CTGCTCCTGGTGACTCTG3'
amplified production length is 582 bases, selects DraI restriction enzyme (Lithuania MBI company) to carry out enzyme and cuts.Enzyme tangent condition is referring to the operational manual of manufacturer.Then on 3% agarose gel, after electrophoresis, with ultraviolet transilluminator, observe length.
result: as shown in Figure 3, while having produced the fragment of two kinds of length of 362 and 220 bases after the long PCR product cutting of 582 bases, the allelotrope that rs605143 site is described is G, when only having 258,220 and illustrate that the allelotrope in rs605143 site is A during 104 3 kind of fragment.Because the testing sample shown in Fig. 3 is the DNA from individuality to be measured, so, as shown in swimming lane 1 and 2, produce two kinds of fragments, the genotype in this individual rs605143 to be measured site is G/G homozygote, carries G allelotrope; As swimming lane 3,4,6 and 7 show, have three fragments, the rs605143 site of this individuality to be measured is A/G heterozygote, carries A, G allelotrope; As shown in swimming lane 5, produce 3 bar segment, the rs605143 site of individuality to be measured is A/A homozygote, carries A allelotrope
embodiment 10, utilize PCR and direct sequencing to detect rs605143 loci polymorphism of the present invention
method: first PCR reaction is carried out in rs605143 site, actual conditions and primer are referring to embodiment 3.PCR product is directly read to sequence on ABI 3500 sequenators.
result: as depicted in figs. 1 and 2, be order-checking process of the present invention and part sequencer map.Wherein in the sequencer map of Fig. 1,1 to 3 show that respectively the rs605143 pleomorphism site of the present invention's individuality to be measured is, GG homozygote, A/G heterozygote and AA homozygote.
embodiment 11, the association study of polymorphic site and coronary heart disease
for further inquiring into the relation between type Ⅳ collagen protein gene α 1 and incidence of coronary heart disease from hereditary angle, and provide the evidence of genetic epidemiology for existing similar research both at home and abroad, use the methods of genotyping of the current international practice of PCR-RFLP, in the 471 routine patients with coronary heart disease of collecting and corresponding 624 example contrasts, vitro detection is from the polymorphism in rs605143 site in the sample of individuality to be measured, thereby analysis rs605143 site is at patients with coronary heart disease and normal control crowd's distributional difference.
first use Hardy-Weinberg balance check.Hardy-Weinberg balance is a kind of concept of population genetics: mainly refer to a group human body that a nationality lives in a certain area, can mutually hybridize at random, whole genetic information that this colony has are called the gene pool of this colony, it directly reflects this area, the hereditary feature that this is national.Expression-form gene frequency and the genotype frequency of these genetic information, do not having under the condition of sudden change, migration and genetic drift, and in colony, gene frequency and genotype frequency are observed to remain unchanged and be Hardy-Weinberg balance.Existing like this genetic polymorphism, possesses again hereditary stability.
gene type interpretation of result is found, gene type result meets Hardy-Weinberg balance, therefore can get rid of experimental error, and this gene type result is reliable, and analytical results is in Table 2.
conclusion: single factor analysis is found, three kinds of genotype individualities of polymorphic site rs605143 are distributed with notable difference (P<0.05) in case group and control group, the aobvious recessive inheritance pattern of A/G is further analyzed discovery, the allelic carrier of the relative A of the allelic carrier of G has marked difference (P<0.05) in case group and control group, wherein the allelic carrier of G to suffer from the danger of coronary heart disease be dangerous 1.4 times that the allelic carrier of A suffers from coronary heart disease.
conclusion: single factor analysis is found, three kinds of genotype individualities of polymorphic site rs605143 are distributed with notable difference (P<0.05) in case group and control group, the aobvious recessive inheritance pattern of A/G is further analyzed discovery, G allelotrope and GG be genotypic be distributed in CHD group apparently higher than control group (distribution of allelotrope G: case group is 0.59, control group is 0.538; The genotypic distribution frequency of GG: CHD group is 37.2, control group is 30.0), and there is statistical significance (P value is all less than 0.05).Above result of study shows, carries risk that G allelotrope person suffers from coronary heart disease higher than carrying allelotrope A person.
disturb the most serious factor of case-control study to comprise crowd's mixing in hereditary basis, when case is selected because the disease that standard does not strictly produce mixes etc., more than having got rid of on the basis of several Confounding Factor, contriver has selected 624 research samples and corresponding contrast, in the international similar research of Zhe, belong to larger research sample, in order further to analyze rs605143 site, be whether independently Hazard Factor and observe the contribution of this site to coronary heart disease risk level of coronary heart disease, adjusting the age, sex, hypertension, diabetes, smoking, triglyceride level, cholesterol, high-density lipoprotein (HDL), after the impact of other Hazard Factor of coronary heart disease such as low-density lipoprotein, carry out multivariate logistic analysis of regression model.Logistic analysis of regression model is a kind of common analytical procedure statistically, and it is mainly used in the analysis of dichotomic variable, can be used for adjusting whole analysis is caused to the factor mixing.The results are shown in Table 3.
result: as shown in table 3, multivariate logistic s analysis of regression model shows, age, sex, hypertension, diabetes, smoking, triglyceride level, cholesterol, high-density lipoprotein (HDL) have been adjusted, after the impact of other Hazard Factor of coronary heart disease such as low-density lipoprotein, find rs605143 site remain coronary heart disease independently Hazard Factor [GG genotype is compared with AA genotype, the risk of suffering from coronary heart disease obviously increases, OR value=1.369,95%CI is (1.021 to 1.834)], and there is statistical significance (P=0.036).
conclusion: the COL4A1 gene that polymorphic site rs605143 sudden change is described is coronary heart disease dependent genes, carrying the individual danger of suffering from coronary heart disease of G allelotrope significantly raises than carrying the allelic individual danger of suffering from coronary heart disease of A, and the change in this site is independently Hazard Factor of coronary heart disease, therefore the test kit of the coronary heart disease dependent genes of rs605143 site mutation clinically can be for detection of, prevention, diagnosis or treatment coronary heart disease.
embodiment 12, and vitro detection coronary heart disease dependent genes is the test kit of COL4A1 gene
a test kit for vitro detection coronary heart disease dependent genes, this test kit comprises:
1) primer in amplification rs605143 site:
upstream primer: 5'AAAGCCATTGCTACCTCA3';
downstream primer: 5'CTGCTCCTGGTGACTCTG3'
2) pcr amplification enzyme and corresponding damping fluid.
) EcoR I restriction enzyme and corresponding restriction enzyme mapping.
?
coronary heart disease dependent genes is the nucleotides sequence list of COL4A1 gene:
Application?Project
<120> Title: the reagent of coronary heart disease dependent genes and vitro detection thereof, preparation or test kit, application
<212>?Type?:?DNA
<211>?Length?:?158,187
sequenceName: coronary heart disease dependent genes is the COL4A1 gene of pleomorphism site rs605143 sudden change
<213>?OrganismName?:?COL4A1
<400>?PreSequenceString?:
aaagccattg?ctacctcagg?acttatcgat?agagcattaa?ccacacaatg
cagccatttg?tgcatgtaag?ttctgccact?gcattcttgt?ctcaagtggg
tacagccagc?tctaaaccat?ctgtggctgc?agataccctt?gatgagaagg
cgttgtaaaa?tgtaaactct?agatttgcac?tttagaactc?ctcagaaata
atcactgcat?aaaatgaatt?taaatggact?ttggtgagga?tgtactattt
tttttttgtt?tgtttgtttt?ttgttttttg?tttttttgtc?agggatttga
agggattctg?gttgagctga?agcaggcagg?agttatgctc?tggaaaagga
gtataacagt?tacatgtctc?tcaatattct?tgagcataca?tttctagagt
caaggaatta?gtatacccta?ggctcacagg?gatgagagat?gtgggttctt
tcagtgaagt?aatttagggg?ctgctctttg?aagcataaac?atggagtcca
ctttgtttaa?gacgatggtg?ccagctgcac?caatgtcgaa?gggctaacga
ggagccagg?caggaccagag?tcaccaggag?cag
Claims (5)
1. coronary heart disease dependent genes is a COL4A1 gene, it is characterized in that the nucleotide sequence of this gene has the sequence shown in SEQ ID NO:1.
2. a reagent for vitro detection coronary heart disease dependent genes according to claim 1, is characterized in that this reagent is for detection of the polymorphism in COL4A1 gene rs605143 site, and this reagent comprises following primer:
Upstream primer: 5'AAAGCCATTGCTACCTCA3'
Downstream primer: 5'CTGCTCCTGGTGACTCTG3'
The reagent of vitro detection coronary heart disease dependent genes according to claim 2, is characterized in that this reagent is for the reagent combining with restriction fragment length polymorphism analysis for polymerase chain reaction or the reagent combining with direct sequencing for polymerase chain reaction.
3. preparation or the test kit of the reagent of the vitro detection coronary heart disease dependent genes described in good grounds claim 2 or 3, is characterized in that this test kit comprises pcr amplification enzyme and corresponding damping fluid.
One kind according to the reagent of the vitro detection coronary heart disease dependent genes described in claim 2 or 3 in the preparation for the preparation of vitro detection coronary atherosclerotic heart disease genes involved or the application in test kit.
5. application according to claim 5, is characterized in that comprising in test kit pcr amplification enzyme and corresponding damping fluid.
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| CN108004313A (en) * | 2017-12-20 | 2018-05-08 | 新疆医科大学第附属医院 | The premature coronary heart disease Disease-causing gene and its reagent of vitro detection, preparation or kit and application |
| CN110205380A (en) * | 2019-07-03 | 2019-09-06 | 贵州省临床检验中心 | One kind SNP marker relevant to cardiovascular disease and its application |
| CN110229882A (en) * | 2019-07-03 | 2019-09-13 | 贵州省临床检验中心 | One kind mthfr gene molecular labeling relevant to cardiovascular disease and its application |
| CN112342293A (en) * | 2020-11-23 | 2021-02-09 | 新疆医科大学第一附属医院 | Family pathogenic coronary heart disease susceptibility gene, reagent for in vitro detection thereof and application thereof |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108004313A (en) * | 2017-12-20 | 2018-05-08 | 新疆医科大学第附属医院 | The premature coronary heart disease Disease-causing gene and its reagent of vitro detection, preparation or kit and application |
| CN108004313B (en) * | 2017-12-20 | 2021-03-16 | 新疆医科大学第一附属医院 | Early-onset coronary heart disease pathogenic gene, reagent, preparation or kit for in vitro detection of early-onset coronary heart disease pathogenic gene and application of early-onset coronary heart disease pathogenic gene |
| CN110205380A (en) * | 2019-07-03 | 2019-09-06 | 贵州省临床检验中心 | One kind SNP marker relevant to cardiovascular disease and its application |
| CN110229882A (en) * | 2019-07-03 | 2019-09-13 | 贵州省临床检验中心 | One kind mthfr gene molecular labeling relevant to cardiovascular disease and its application |
| CN112342293A (en) * | 2020-11-23 | 2021-02-09 | 新疆医科大学第一附属医院 | Family pathogenic coronary heart disease susceptibility gene, reagent for in vitro detection thereof and application thereof |
| CN112342293B (en) * | 2020-11-23 | 2024-02-06 | 新疆医科大学第一附属医院 | Family pathogenic coronary heart disease susceptibility gene, reagent for in vitro detection and application thereof |
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| CN103667301B (en) | 2016-09-14 |
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