CN103160592A - Molecular biology method for predicating endurance potentials of excellent ice-snow sportsmen - Google Patents
Molecular biology method for predicating endurance potentials of excellent ice-snow sportsmen Download PDFInfo
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Abstract
The invention discloses a molecular biology method for predicating the endurance potentials of excellent ice-snow sportsmen, which can be used for inquiring the relationship between the I/D (kininase II gene insertion/deletion) polymorphisms of ACE (angiotensin-converting enzyme) and the endurance potentials of the excellent ice-snow sportsmen. Lots of research results indicate that the genetic I/D polymorphisms are associated with the aerobic endurance of the sportsmen, are the key factors for judging the aerobic endurance quality of a human body, and affect the optimal level of the aerobic endurance quality of a human body and the sensitivity of a human body for endurance training; and the genes mainly affect the cardio-pulmonary function of a human body, and thus affecting the aerobic endurance quality of a human body. In consideration that the ACE genes are related to the aerobic endurance of movements, researches for the distribution of the genes in the excellent ice-snow sportsmen by applying the molecular biology method have important theoretical significance for determining the relevance between the genes and endurance-type ice-snow sports, and lay the foundation of future molecular selection by applying the molecular biology method.
Description
Technical field
The present invention relates to a kind of molecular biology method of predicting outstanding winter sports person's endurance potential, be exactly specifically the polymorphism by analysis ACE (Angiotensin-converting enzyme) I/D (kininase II gene insertion/deletion type), thereby learn that the ACE gene pleiomorphism is by the talent of the outstanding endurance of which kind of approach impact.This invention belongs to the interdisciplinary field of Basic Life science, mankind's inheritance resources, molecular biology and sports science.
Background technology
ACE is 20 peptide strand acid glycoproteins, for containing the metallohydrolases of zinc, its system peptidyl one or two peptidohydrolases by name or kininase II.ACE extensively is present in each tissue of whole body, mainly is distributed in vascular endothelial cell (especially common with pulmonary circulation), renal epithelial cell etc.Bradykinin is the natural substrate of ACE, suppresses its biologic activity by two C-terminal dipeptides of cracking.
Renin-angiotensin system (Renin-anglotensin system, RAS) comprise proangiotensin (Anglotensin origin, be abbreviated as AO), angiotensin I (Anglotensin I, be abbreviated as Ang I), Angiotensin II (Anglotensin II, be abbreviated as Ang II), feritin, ACE and angiotensin-ii receptor (Anglotensin II receptor is abbreviated as AT).RAS acts in cardiovascular systems significantly, as adjusting blood pressure, Q volume of blood, antiotasis, and causes vessel wall thickening or myocardial cell's hypertrophy and non-myocardial cell's propagation.Wherein Ang II is the effector peptide in RAS, and AT causes cardiac muscle and vasoconstriction by its acceptor, regulates electrolyte balance, causes hyperplasia, propagation etc.Ang II has direct and of short duration effect to heart, and comprising increases heart rate and strengthen cardiac contractility; Simultaneously, Ang II also has the effect of its structure of long-term effect to heart, and it mainly stimulates collagen and sheath to generate by hypertrophy and the inoblast that causes the myocardial cell.
ACE is the key enzyme in RAS enzyme/substrate chain reaction system, and the effect of ACE is that catalysis decapeptide Ang I changes octapeptide Ang II into.The ACE level is determining Ang II growing amount in RAS.Numerous studies confirm that, the ACE horizontal stable, measuring repeatability is good, and environment metabolism and Hormone Factors are less on its impact.But research confirms that also there is obvious mutability in the ACE level between individuality, and difference can reach 5 times more than.Obviously, the ACE level is stablized on the one hand with in individuality, and is associated with lacking environmental factors, on the other hand due to the larger phenomenon of interindividual variation, makes people infer that the ACE level is largely by genetic determination.
The ACE gene is positioned at long-armed 2 district's 3 bands (17q23) of karyomit(e) No. 17.ACE gene span is 21kb, and 26 exons and 25 introns are arranged.Found at present 6 kinds of ACE gene polymorphic signs, but the Alu Insert Fragment of the 287bp that exists in the 16th intron and/or deletion fragment most study.Thus, the ACE gene can be divided into DD homozygous deletion type, the insertion of ID heterozygote or absence type and three kinds of different genotype of II homozygote insert type in this polymorphic sign site.DNA polymorphism refers to the otherness that in karyomit(e) allelotrope, Nucleotide is arranged.There is two or more form in allelotrope in the DNA zone (or fragment), can be divided into sequence polymorphism and sequence-length polymorphism.Wherein sequence-length polymorphism is connected with core sequence separately by tumor-necrosis factor glycoproteins and is distributed on karyomit(e) with repeating or be dispersed in, form polymorphism (as Alu repeated sequence) in the insertion/deletion mode, this polymorphism is the genetic marker of present most worthy.
The influence factor of maximum aerobic capacity can be summarized in central mechanism and periphery mechanism.Wherein, ventricular volume and stroke output are the core links in central mechanism, and the conformability of good cardiovascular function, particularly artery and heart is the key of outstanding endurance exercise person's success; Muscle fiber types, flesh capillary density and oxidasic activity are the dominant roles in periphery mechanism.ACE gene I/D polymorphism and sportsmen's aerobic endurance exist related, are the key factors of decision human body aerobic stamina diathesis, affect simultaneously the optimum level of human body aerobic stamina diathesis and human body to the susceptibility of endurance training.The cardio-pulmonary function of this gene major effect human body, thus the aerobic stamina diathesis of human body affected.
Summary of the invention
The present invention can predict outstanding winter sports person's endurance potential.
The purpose of this invention is to provide a kind of molecular biology method of predicting outstanding winter sports person's endurance potential.
Forecasting Methodology provided by the present invention is ACE gene I/D (kininase II gene insertion/deletion type) polymorphism analysis.Be positioned at the insertion of ACE gene intron 16 or deletion polymorphism after PCR method detects, can present 3 kinds of banding patterns: what the 401bp band was only arranged is homozygote insert type (II), what the 113bp band was only arranged is homozygous deletion type (DD), and what 401bp and two bands of 113bp were arranged is that heterozygote inserts or absence type (I/D).Between DD, ID, II genotype levels of serum ACE three, there were significant differences, and DD type people's ACE level is the highest, and the ID type takes second place, and II type people is minimum.
Embodiment
The below carries out ACE gene I/D (kininase II gene insertion/deletion type) polymorphism analysis take outstanding winter sports person and common artificial experimental subjects and describes concrete enforcement of the present invention in detail as example.
Embodiment: ACE gene I/D (kininase II gene insertion/deletion type) polymorphism analysis
Research object
Research object is mainly divided 2 groups, outstanding winter sports person's group and the general population's group, and the general population organizes main selection people from this area suitable with this elite's age, carries out in strict accordance with the principle of stochastic sampling, all without professional training history.
The mensuration of ACE gene I/D polymorphism
(1) extraction of DNA
Get the 2ml whole blood from ulnar vein and inject heparin (10U/ml) anti-freezing test tube, then centrifugal (1500rpm, 10min) immediately.After the red corpuscle of dissolving in whole blood, go in 1.5ml eppendorf pipe, be prepared into the white corpuscle precipitation be stored in-80 ℃ standby.Use classical approach DNA extraction agent box total DNA of extracting from the white corpuscle precipitation.
(2) design of primers and PCR product order-checking
Design of primers first finds the ACE gene order of publishing from the Genbank database by the internet, select suitable sequence scope, adopt Primer select5.0 primer-design software, the general long 21～24bp of the primer of designing, annealing temperature is 58 ℃～60 ℃, by having obtained corresponding band after pcr amplification.Amplified production is carried out nucleotide sequencing, confirm that amplified production sequence and bibliographical information are in full accord, illustrate that the selected gene of above-mentioned primer pair has high degree of specificity.
The sequence of primer is as follows:
Upper primer (Forward): 5 ' TCC CAT TTC TCT AGA CCT GCT G3 '
Lower primer (Reverse): 5 ' GCC CTT AGC TCA CCT CTG CT3 '
(3) PCR reaction system (25l)
The total reaction system is 25l.H
2O12.8l、5×buffer5l、10mM?dNTPs0.75l、250mM?Mg
2+0.4l、10M?primer?for0.4l、10M?primer?Rev0.4l、Ext?aq?Hs0.25l、DNA(10ng/l)5l。
(4) PCR reaction conditions
95 ℃ of initial stage denaturations, are extended 72 ℃ of 60s at 5min, 95 ℃ of 30s of sex change, 58 ℃ of 30s of annealing, 2-4 * 40cycles, extend 72 ℃ of 7min, 4 ℃ of 10min at last.
(5) electrophoresis
PCR reaction product 10l+6 * loading buffer2l in 1.6%Agarose gel electrophoresis (glue 45ml+FB2l) imaging, is developed under ultraviolet lamp, take a picture and image analysis system is taken the photograph sheet by gel, and definite genotype.
(6) the genotypic judgement of ACE and qualification result
Be positioned at the insertion of ACE gene intron 16 or deletion polymorphism after above-mentioned PCR method detects, can amplify the fragment of two kinds of length, it is the deletion fragment (D) that 401bp inserts amplified fragments (I) and 113bp, can present following 3 kinds of banding patterns under ultraviolet lamp: what the 401bp band was only arranged is homozygote insert type (II), what the 113bp band was only arranged is homozygous deletion type (DD), and what 401bp and two bands of 113bp were arranged is that heterozygote inserts or absence type (I/D).
The insertion/deletion (I/D) that studies show that the Alu repeated sequence of the one section 287bp that is positioned at the 16th intron has polymorphism, makes the ACE gene that 3 kinds of genotype: II (inserting homozygous), ID (insertion/deletion type) and DD (lacking homozygous) be arranged.ACE is affected by inherited genetic factors at blood level, confirm that the ACE gene pleiomorphism has very large related with blood plasma and the interior ACE level of cell.Between DD, ID, II genotype levels of serum ACE three, there were significant differences, and DD type people's ACE level is the highest, and the ID type takes second place, and II type people is minimum.
ACE gene I/D polymorphism and sportsmen's aerobic endurance exist related, are the key factors of decision human body aerobic stamina diathesis, affect simultaneously the optimum level of human body aerobic stamina diathesis and human body to the susceptibility of endurance training.The cardio-pulmonary function of this gene major effect human body, thus the aerobic stamina diathesis of human body affected.
Claims (3)
1. method of probing into ACE (Angiotensin-converting enzyme) I/D (kininase II gene insertion/deletion type) polymorphism and outstanding winter sports person's endurance potential relation.
2. an insertion homozygous (II) gene, insertion/deletion type (ID) gene and homozygous (DD) gene cloning of disappearance and detection method.
3. method according to claim 1, the innovative point of this method is the dependency by quantitative evaluation ACE gene I/D polymorphism and outstanding winter sports person's endurance, thereby can be with ACE gene I/D polymorphism as one of candidate locus of predicting the selection of outstanding winter sports person's endurance potential gene.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2018036495A1 (en) * | 2016-08-24 | 2018-03-01 | 厦门美因生物科技有限公司 | Qpcr genotyping technique-based athleticism gene detection and evaluation method and system |
| CN113205883A (en) * | 2021-04-12 | 2021-08-03 | 中国人民解放军陆军军医大学第二附属医院 | Method and system for predicting cardiopulmonary function from genetic information and post-exercise physiological parameters |
| CN113430275A (en) * | 2021-06-25 | 2021-09-24 | 北京体育大学 | Method, primer set and kit for screening athletes by adopting LPL gene rs283 polymorphic site |
-
2013
- 2013-04-03 CN CN201310113515.8A patent/CN103160592B/en not_active Expired - Fee Related
Non-Patent Citations (2)
| Title |
|---|
| 杨贤罡等: "ACE基因I/D多态性与耐力素质的关联性研究:Meta分析", <体育科学> * |
| 杨震等: "力量素质相关基因多态性的研究进展", 《西安体育学院学报》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2018036495A1 (en) * | 2016-08-24 | 2018-03-01 | 厦门美因生物科技有限公司 | Qpcr genotyping technique-based athleticism gene detection and evaluation method and system |
| CN113205883A (en) * | 2021-04-12 | 2021-08-03 | 中国人民解放军陆军军医大学第二附属医院 | Method and system for predicting cardiopulmonary function from genetic information and post-exercise physiological parameters |
| CN113430275A (en) * | 2021-06-25 | 2021-09-24 | 北京体育大学 | Method, primer set and kit for screening athletes by adopting LPL gene rs283 polymorphic site |
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