CN103966207A - New short nucleotide tandem repeat sequence locus and application thereof - Google Patents
New short nucleotide tandem repeat sequence locus and application thereof Download PDFInfo
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Abstract
The invention discloses a new short nucleotide tandem repeat sequence locus and application thereof. The invention provides the short tandem repeat locus G2S0002, the sequence of which is shown as SEQ ID NO.:1, and the short tandem repeat locus G2S0002 can be used for preparation of: (a) reagents or kits for genetic relationship analysis; (b) reagents or kits for individual identification; (c) reagents or kits for paternity test or blood relationship analysis; (d) reagents or kits for detecting whether maternal blood contamination exists in extracted amniotic fluid; and/or (e) kits for detecting whether white blood cells in a bone marrow transplantation receptor is replaced by donor cells. The short tandem repeat locus G2S0002 has a high degree of distinction, and can effectively analyze the genetic relationship.
Description
Technical field
The present invention relates to a kind of short Nucleotide tandem repetitive sequence site and application thereof.
Background technology
STR (Short Tandem Repeat.STR), claim again microsatellite DNA (microsatellite DNA), a 2-6bp of class repeating unit, multiplicity 10-60, the strand of dna connection tumor-necrosis factor glycoproteins of fragment length below 400bp; Its produce reason be to slide in DNA replication dna process, or in the time copying and repair sliding chain and complementary strand base mispairing, cause disappearance or the insertion of one or several repeating unit.That STR has is widely distributed in genome, allelotrope is many, heterozygosity is high, somatotype result is stable, STR heredity meets Meng get Er law of inheritance (Koreth, J., et al.1996.Microsatellites andPCR ge-nomic analysis.J.Pathol.178:239-248.) etc. feature, and when multiple str locus seat joint-detection, individual recognition power and parentage exclusion probability are high.Thereby STR is with features such as its genetic polymorphisms, be used as s-generation genetic marker and be widely used in the fields such as medical science individual recognition, the drafting of human inheritance's collection of illustrative plates, paternity test, legal medical material evidence examination, effectively promoted forensic identification and changed to individual recognition from the individuality eliminating of material evidence sample in the past comprehensively.
At present, the technology of individual recognition mainly adopts STR-PCR to detect, to multiple str locus seat united typings, to determine parental right relation or individual recognition etc., all relatively ripe at experimental technique, but in the selection of str locus seat, not unified standard.In the detection kit that manufacturer provides both at home and abroad, site used is not quite similar.Early stage America and Europe selects 10 str locus seats to add 1 sex determining gene (E.A.Cotton, R.F.Allsop, J.F.Guest, R.R.E.Frazier, P.Koumi, I.P.Callow, A.Seager, R.L.Sparkes, Validation of the
sGM Plus
tMsystem for use inforensic casework, Forensic Sci.Int.112 (2000) 151 – 161.).Along with technical development, database increase, comparison information difficulty strengthened afterwards, and the selection of str locus seat is also being updated.More influential database CODIS (Combined DNA Index System) is based on 13 str locus seat (D3S1358 by U.S. FBI now, vWA, D16S539, D5S818, D7S820, CSF1PO, D13S317, TPOX, D8S1179, D21S11, D18S51, TH01, FGA.) build, thereby present most of STR detection kit manufacturer also increases or deletes that some sites are to improve detection efficiency and accuracy rate (D.J.French on the basis of these 13 core gene seats, R.L.Howard, N.Gale, T.Brown, D.G.McDowell, P.G.Debenhan, Interrogation of short tandem repeats usingfluorescent probes and melting curve analysis:A step towards rapid DNAidentity screening.Forensic Science. (2008) 333-339).For example Promega company
16System test kit has comprised 13 core gene seats, and has increased PentaD, PentaE and tri-locus of AMELO; American AB I company
pCRAmplification Kit is that 13 core gene seats add D2S1338, D19S433 and AMELO; Other two test kit of its company
sGM Plus
tMpCR Amplification Kit and
pCR Amplification Kit has also done the increase and decrease of corresponding gene seat point.The unit of the similar detection kit of domestic production also has a lot, the DNA Typer that for example Ministry of Public Security two promotes
tM15, middle dolantin joins the AGCU EX22STR fluorescence detection reagent kit of Bioisystech Co., Ltd, and the locus that detects increases to 22.
The advantage of the str locus seat that above test kit is selected is, taking 13 core gene seats that gain public acceptance as basis, the quantity of information obtaining is detected in more additional other sites to enrich, can more comprehensively reflect the otherness between individuality, and then complete the work such as individual recognition, patriarchy judgement.But be not all optimal selection in these locus used, for example FGA locus exists that sequence length is long, repeating unit is many but span is discontinuous, thereby affect and the compatibility in other site, be unfavorable for using with the multiple system of multiple locus joint-detection; D19S433 locus is positioned at genome camber tumor-necrosis factor glycoproteins district, and amplimer design exists compared with burden, and the stability in this site is not high, and the available information quality providing while being applied to individual recognition and discrimination are not high; For another example D21S11 locus is positioned at the problem in genome camber tumor-necrosis factor glycoproteins region except existing, and also finds may have the CNV (situation of gene copy number variation in this locus region; In the time running into the improper pattern detection of gene copy number, can reduce the accuracy of this site STR somatotype, in individual's identification, adopt the effect of this locus also can have a greatly reduced quality.Along with the foundation of human genome collection of illustrative plates and the development of sequencing technologies, with lower cost, within the time faster, the full genome of the individual order of resurveying is become to possibility, and then can under larger quantity of information, filter out the str locus seat that discrimination is higher, promote to be applied to selection and the renewal of the str locus seat in the researchs such as individual recognition.
Therefore, for overcoming poor compatibility, have CNV or being positioned at the defects such as highly repetitive sequence region, this area a kind of novel STR with high discrimination of still needing.
Summary of the invention
The object of the present invention is to provide a kind of novel STR locus, do not have CNV, and be not positioned at highly repetitive sequence region, there is high discrimination.
A first aspect of the present invention, provides a kind of STR locus G2S0002 of separation, and sequence is as shown in SEQ ID NO.:1.
A second aspect of the present invention, provides a kind of nucleotide sequence of separation, and the structure of described nucleotide sequence is as follows:
X1-X2-X3
In formula, X1 is 1-782 position in SEQ ID NO.:2;
X2 is the genetic polymorphism district of contain >=2 repeating unit aaag;
X3 is 873-1652 position in SEQ ID NO.:2.
A third aspect of the present invention, provides the purposes of the STR locus G2S0002 described in first aspect, for the preparation of:
(a) reagent or the test kit analyzed for genetic affinity;
(b) for reagent or the test kit of individual recognition;
(c) for reagent or the test kit of paternity test or consanguinity analysis;
(d) for detection of the reagent or the test kit that whether exist female blood stains to dye in extracting amniotic fluid; And/or
(e) test kit whether being replaced by donorcells for detection of the white corpuscle in acceptor after bone marrow transplantation.
In another preference, described reagent comprises (i) primer or primer pair; (ii) probe; (iii) nucleic acid chip.
In another preference, the sequence of described primer pair is as shown in SEQ ID NO.:3 and SEQ ID NO.:4.
A fourth aspect of the present invention, provides the primer pair of a kind of specific amplification STR locus G2S0002, and the sequence of described primer pair is as shown in SEQ ID NO.:3 and SEQ ID NO.:4.
In another preference, described STR locus G2S0002 is as described in first aspect.
A fifth aspect of the present invention, provides a kind of test kit, comprising:
(a) for the primer pair of specific amplification STR locus G2S0002;
(b) working instructions.
In another preference, described STR locus G2S0002 is as described in first aspect.
In another preference, described test kit also comprises that (c) specific amplification is selected from the primer pair of one or more locus of lower group:
D3S1358、vWA、D16S539、D5S818、D7S820、CSF1PO、D13S317、TPOX、D8S1179、D21S11、D18S51、TH01、FGA、D2S1338、D19S433、AMELO。
A sixth aspect of the present invention, provides a kind of amplification system, comprising:
(a) for the primer pair of specific amplification STR locus G2S0002; With
(b) optional, specific amplification is selected from the primer pair of one or more locus of lower group:
D3S1358、vWA、D16S539、D5S818、D7S820、CSF1PO、D13S317、TPOX、D8S1179、D21S11、D18S51、TH01、FGA、D2S1338、D19S433、AMELO；
And described primer pair (a) and optional primer pair (b) are positioned at same amplification system.
In another preference, described STR locus G2S0002 is as described in first aspect.
In another preference, described amplification system is composite amplification system, and wherein, described primer pair (b) comprises 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15, or 16 pairs of Auele Specific Primers pair; And/or
Described composite amplification system is fluorescently-labeled composite amplification system, wherein, described fluorescent mark refers to the primer of locus described in one or more fluorescent labeling reagent marks of selecting lower group: FAM, HEX, VIC, NED, PET, TAMRA, ROX, fluorescein, FITC, IRD-700/800, CY3, CY5, CY3.5, CY5.5, TET, TAMRA, JOE, BODIPY TMR, Oregon is green, rhodamine is green, rhodamine is red, texas Red or Ya Jima Huang.
A seventh aspect of the present invention, provides a kind of amplification method, comprises step:
In amplification system described in aspect the 6th, taking sample to be detected as template, utilize polymerase chain reaction to carry out the amplification of locus, thereby obtain locus amplified production.
In another preference, described method also comprises: the step that described locus amplified production is detected.
In another preference, described detection is selected from lower group: capillary electrophoresis, order-checking, hybridization.
In another preference, described testing sample is selected from lower group: blood, saliva, seminal fluid, sweat, amniotic fluid, bone or hair.
A eighth aspect of the present invention, provides a kind of genetic affinity analytical procedure, comprises step:
In amplification system described in aspect the 6th, taking sample to be detected as template, utilize polymerase chain reaction to carry out the amplification of locus, thereby obtain locus amplified production; With
Wherein, described sample to be detected comprises the sample of nucleic acid S1 from Different Individual respectively, S2 ..., Si, the positive integer that in formula, i is >=2;
(2) to from different IPs acid sample S1, S2 ..., the locus amplified production of Si detects, thereby obtains the detected result corresponding to different IPs acid sample;
(3) detected result step (2) being obtained compares, thereby determines the genetic affinity between Different Individual.
In another preference, described genetic affinity analysis comprises individual recognition analysis, paternity test analysis, genetic connection analysis.
The invention provides a kind of novel str locus seat, there is not CNV, and be not positioned at highly repetitive sequence region, heterozygosity is high, resolving power is high, compatible high, can in same system, use with existing most of sites, can effectively distinguish random crowd's genetic affinity.
In should be understood that within the scope of the present invention, above-mentioned each technical characterictic of the present invention and can combining mutually between specifically described each technical characterictic in below (eg embodiment), thus form new or preferred technical scheme.As space is limited, tire out and state no longer one by one at this.
Brief description of the drawings
Fig. 1 is the detected peaks figure in the multiple STR of male parent site.
Fig. 2 is the detected peaks figure in maternal multiple STR site.
Fig. 3 is the detected peaks figure in the multiple STR of filial generation site.
Fig. 4 is the detected peaks figure in the multiple STR of affinity-less relation sample site.
Embodiment
The inventor, through extensive and deep research, by a large amount of screenings, filters out a kind of novel STR locus G2S0002, the genetic polymorphism district of contain >=2 repeating unit aaag first.This STR locus discrimination reaches more than 0.9197, with the str locus seat coupling of a small amount of routine, can effectively distinguish genetic affinity, and discrimination is up to more than 0.99.On this basis, completed the present invention.
STR locus
The present invention, by a large amount of screenings, isolates a kind of new STR locus, and STR site information is as shown in table 1.
The essential information of table 1STR
| Site title | Repeat number | Repeating unit | Chromosome position | Initial | Finish |
| G2S0002 | 22.2 | aaag | 2 | 142567665 | 142567754 |
The STR locus of a kind of separation provided by the invention, called after G2S0002, sequence is as shown in SEQ ID NO.:1.
Repeat number 22.2,22 repeating units add two base aa, and wherein the aagg in SEQ ID NO.:1 can be used as another kind of repeat unit, therefore also contributes in repeat number.
The nucleotide sequence of separation provided by the invention, structure is as follows:
X1-X2-X3
In formula, X1 is 1-782 position in SEQ ID NO.:2;
X2 is the genetic polymorphism district of contain >=2 repeating unit aaag;
X3 is 873-1652 position in SEQ ID NO.:2.
In another preference, the length of described X2 is 50-150bp, preferably 60-130bp or 70-120bp, more preferably 80-110bp.
In another preference, described X2 is the genetic polymorphism district of contain >=10 repeating unit aaag.
In another preference, described X2 is the genetic polymorphism district of contain >=20 repeating unit aaag.
In another preference, X2 is the polynucleotide shown in SEQ ID NO.:1 or its fragment (being included in the fragment that 5' end, 3' end and/or region intermediate form because lacking one or more bases).
STR locus G2S0002 of the present invention, can be used in:
(a) reagent or the test kit analyzed for the preparation of genetic affinity;
(b) for the preparation of reagent or the test kit of individual recognition;
(c) for the preparation of reagent or the test kit of paternity test or consanguinity analysis;
(d) for the preparation of detecting the reagent or the test kit that whether exist female blood stains to dye in extracting amniotic fluid; And/or
(e) test kit whether being replaced by donorcells for the preparation of the white corpuscle in acceptor after detection bone marrow transplantation.
Described reagent also comprises (i) primer or primer pair; (ii) probe; (iii) nucleic acid chip.Preferably, the sequence of described primer pair is as shown in SEQ ID NO.:3 and SEQ ID NO.:4.
The present invention also provides the primer pair of a kind of specific amplification STR locus G2S0002, and sequence is as shown in SEQ ID NO.:3 and SEQ ID NO.:4.
One or two primer in described primer pair is with detectable.
Described detectable comprises: fluorescent marker or quantum dot-labeled thing.
Described fluorescent mark includes but not limited to: FAM, HEX, VIC, NED, PET, TAMRA, ROX, fluorescein, FITC, IRD-700/800, CY3, CY5, CY3.5, CY5.5, TET, TAMRA, JOE, BODIPY TMR, Oregon is green, rhodamine is green, rhodamine is red, texas Red or Ya Jima Huang.The marker that above-mentioned fluorescent marker is known in the art, all has commercial goods, can obtain by purchase.
Test kit
Test kit provided by the invention, comprising:
(a) for the primer pair of specific amplification STR locus G2S0002;
(b) working instructions.
Preferably, the sequence of described primer pair is as shown in SEQ ID NO.:3 and SEQ ID NO.:4.
Described test kit also comprises that (c) specific amplification is selected from the primer pair of one or more locus of lower group:
D3S1358、vWA、D16S539、D5S818、D7S820、CSF1PO、D13S317、TPOX、D8S1179、D21S11、D18S51、TH01、FGA、D2S1338、D19S433、AMELO。
In described test kit, described each Auele Specific Primer is to being arranged in same or different containers.
In another preference, described test kit also comprises one or more following reagent:
(d) for the reagent of pcr amplification;
(e) for the reagent of electrophoresis;
(f) for the reagent of extracting DNA;
(g) standard substance.
Amplification system and amplification method
Amplification system provided by the invention, comprising:
(a) for the primer pair of specific amplification STR locus G2S0002; With
(b) optional, specific amplification is selected from the primer pair of one or more locus of lower group:
D3S1358、vWA、D16S539、D5S818、D7S820、CSF1PO、D13S317、TPOX、D8S1179、D21S11、D18S51、TH01、FGA、D2S1338、D19S433、AMELO；
And described primer pair (a) and optional primer pair (b) are positioned at same amplification system.
Preferably, the sequence of described primer pair is as shown in SEQ ID NO.:3 and SEQ ID NO.:4.
In another preference, described amplification system is polymerase chain reaction PCR amplification system.
In another preference, described amplification system is liquid phase.
In another preference, described amplification system contains the reagent for PCR polymerization such as polysaccharase, dNTP.
Preferably, described amplification system is composite amplification system, and wherein, described primer pair (b) comprises 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15, or 16 pairs of Auele Specific Primers pair.
Preferably, described composite amplification system is fluorescently-labeled composite amplification system, wherein, described fluorescent mark refers to the primer of locus described in one or more fluorescent labeling reagent marks of selecting lower group: FAM, HEX, VIC, NED, PET, TAMRA, ROX, fluorescein, FITC, IRD-700/800, CY3, CY5, CY3.5, CY5.5, TET, TAMRA, JOE, BODIPY TMR, Oregon is green, rhodamine is green, rhodamine is red, texas Red or Ya Jima Huang.
In another preference, described each to a primer in primer 5 ' end with fluorescent mark.
Amplification method of the present invention, comprises step:
In above-mentioned amplification system, taking sample to be detected as template, utilize polymerase chain reaction to carry out the amplification of locus, thereby obtain locus amplified production.
Described method also comprises: the step that described locus amplified production is detected.Preferably, described detection adopts the method that is selected from lower group: capillary electrophoresis, order-checking, hybridization.
Described testing sample is selected from lower group: blood, saliva, seminal fluid, sweat, amniotic fluid, bone or hair.
In another preference, described sample comes from people.
Genetic affinity analytical procedure
The invention provides a kind of genetic affinity analytical procedure, comprise step:
(1), in above-mentioned amplification system, taking sample to be detected as template, utilize polymerase chain reaction to carry out the amplification of locus, thereby obtain locus amplified production; With
Wherein, described sample to be detected comprises the sample of nucleic acid S1 from Different Individual respectively, S2 ..., Si, the positive integer that in formula, i is >=2;
(2) to from different IPs acid sample S1, S2 ..., the locus amplified production of Si detects, thereby obtains the detected result corresponding to different IPs acid sample;
(3) detected result step (2) being obtained compares, thereby determines the genetic affinity between Different Individual.
In another preference, described genetic affinity analysis comprises individual recognition analysis, paternity test analysis, genetic connection analysis.
In another preference, described Different Individual is from same family.
The above-mentioned feature that the present invention mentions, or the feature that embodiment mentions can arbitrary combination.All features that this case specification sheets discloses can with any composition forms use, each feature disclosing in specification sheets, can anyly provide the alternative characteristics of identical, impartial or similar object to replace.Therefore apart from special instruction, the feature disclosing is only the general example of equalization or similar features.
Usefulness of the present invention is:
(1) provide a kind of novel str locus seat, do not had CNV, and be not positioned at highly repetitive sequence region.
(2) heterozygosity of new str locus seat is high, and resolving power is high.
(3) new str locus seat compatibility is high, can in same system, use with existing most of sites.
(4) new str locus seat or be combined with known str locus seat, can effectively distinguish random crowd's genetic affinity.
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment are only not used in and limit the scope of the invention for the present invention is described.The experimental technique of unreceipted actual conditions in the following example, conventionally according to normal condition, the people such as such as Sambrook, molecular cloning: laboratory manual (New York:Cold Spring Harbor Laboratory Press, 1989) condition described in, or the condition of advising according to manufacturer.Unless otherwise indicated, otherwise per-cent and umber are weight percent and parts by weight.
Unless otherwise defined, the same meaning that all specialties that use in literary composition and scientific words and one skilled in the art are familiar.In addition, any method similar or impartial to described content and material all can be applicable in the inventive method.The use that better implementation method described in literary composition and material only present a demonstration.
Embodiment 1
By str locus seat G2S0002 new the present invention and 10 conventional locus sites (VWA, D16S539, D5S818, D7S820, D13S317, D8S1179, D18S51, TH01, D2S1338, AMELO) combination, adopt multiple fluorescence PCR technical inspection sample, 4 samples used are respectively akin family of three sample, and the check sample of 1 affinity-less relation.G2S0002 and 10 conventional locus primer sequences are as shown in table 2.
Table 2 primer sequence
Note: PET, VIC, NED, tetra-kinds of fluorescence dyes of FAM ((Applied Biosystems, USA company).
Specific experiment operation steps is as follows:
1) DNA sample is prepared
4 individual of samples of choosing are gathered to blood; Obtained DNA sample and respectively got 1 μ l by the extracting of DNA extraction test kit, its sample is carried out to quality inspection to 1%agarose electrophoresis and concentration is estimated, then according to the concentration of estimating, sample is diluted to working concentration 5-10ng/ μ l.
2) PCR reaction
Respectively get 1ul sample, carry out PCR reaction.
PCR reaction system cumulative volume is 10 microlitres (1 μ L10 × PCR damping fluid (Qiagen company), 0.2 μ L25mmol magnesium chloride, 1 μ L detect mix primer, 0.8 μ L2.5mmol dNTP, 0.04 μ L HotStarplus Taq enzyme (Qiagen company), the 5.96 μ L ddH2O of multiple str locus seats).
PCR reaction conditions arranges as follows: 95 DEG C 10 minutes; The Touchdown program of 7 circulations (94 DEG C 20 seconds, 65 DEG C-1 DEG C/circulation 40 seconds and 72 DEG C of 2min), 28 cyclic amplifications (94 DEG C 20 seconds, 63 DEG C of 30 seconds and 72 DEG C of 2min), extend 2 minutes at 72 DEG C, extend 60 minutes 4 DEG C of preservations at 60 DEG C;
After PCR has reacted, by 20 times of pcr amplification product dilutions, take out 1ul and add 8.9ul Hi-Di(height deionized formamide) and 0.1ul LIZ(ABI house journal dyestuff, be used to mark in tagged molecule amount), mix rear 95 DEG C of 5min, capillary electrophoresis loading; Detect primer with fluorescence because of STR used site, detect the amplification segment of different colours fluorescence and different sizes by capillary electrophoresis, draw STR somatotype information with fluorescence detecting system analytical results, result is as shown in Fig. 1-4 and table 3.
The gene type result of 11 locus is as shown in table 3; Sample 1 represents male parent, and sample 2 represents maternal, and sample 3 represents filial generation, and sample 4 represents check sample.
Table 3 gene type result
Note: in form, X and Y represent sex chromosome, digital represent the relative repetition numerical value of repeating unit in sample allelotrope.
Result shows, the G2S0002 site that adopts new screening to obtain obviously can verify between sample 1 and sample 2, sample 3 sibship, with sample 4 affinity-less relations, the G2S0002 site that new screening obtains has shown higher discrimination in result, be applied in the biomedical detection such as STR somatotype, paternity identification, individual recognition, can effectively improve somatotype recognition efficiency and accuracy, and G2S0002 locus site has good compatibility, energy and most existing str locus seat point are used in combination, and have very strong practicality.
Embodiment 2
Choose at random 20 groups of samples with sibship and affinity-less relation, adopt the method for embodiment 1 to detect, difference is, only adopts the new str locus seat G2S0002 of the present invention.
After testing, after definite male parent sample, then determine in remaining 19 groups of samples and the akin filial generation sample of male parent, the new str locus seat G2S0002 of the present invention can accurately get rid of the wherein sample of 12 groups of affinity-less relations, has shown higher discrimination.
Adopt the method for embodiment 1, by str locus seat G2S0002 new the present invention and 6 conventional locus sites (VWA, D16S539, D18S51, TH01, D2S1338, AMELO) combination, remaining 7 groups of samples are detected, precise Identification its sibship.
The gene type result of random 20 affinity-less relations of table 4 and akin sample
Note: 1. sample 1 represents male parent, sample 2 represents maternal, sample 3 represents filial generation, other sample affinity-less relation; 2. for determining that sex adds AMELO; 3. in form, X and Y represent sex chromosome, digital represent the relative repetition numerical value of repeating unit in sample allelotrope.
Result shows, only, by str locus seat G2S0002 of the present invention, just can provide extremely valuable somatotype information.Certainly, being combined in the site that G2S0002 is known with other to provide somatotype information more accurately, for example, additional two the site D18S51 of G2S0002 and D16S539, just can accurately get rid of the sample of affinity-less relation in remaining 7 groups, additional more site, accuracy is higher.In existing conventional Sites Combination, add G2S0002 as seen, can greatly improve the accuracy of individual recognition.
All documents of mentioning in the present invention are all quoted as a reference in this application, are just quoted separately as a reference as each section of document.In addition should be understood that those skilled in the art can make various changes or modifications the present invention after having read above-mentioned teachings of the present invention, these equivalent form of values fall within the application's appended claims limited range equally.
Claims (12)
1. a STR locus G2S0002 for separation, is characterized in that, the sequence of described locus is as shown in SEQ ID NO.:1.
2. a nucleotide sequence for separation, is characterized in that, the structure of described nucleotide sequence is as follows:
X1-X2-X3
In formula, X1 is 1-782 position in SEQ ID NO.:2;
X2 is the genetic polymorphism district of contain >=2 repeating unit aaag;
X3 is 873-1652 position in SEQ ID NO.:2.
3. the purposes of STR locus G2S0002 as claimed in claim 1, is characterized in that, for the preparation of:
(a) reagent or the test kit analyzed for genetic affinity;
(b) for reagent or the test kit of individual recognition;
(c) for reagent or the test kit of paternity test or consanguinity analysis;
(d) for detection of the reagent or the test kit that whether exist female blood stains to dye in extracting amniotic fluid; And/or
(e) test kit whether being replaced by donorcells for detection of the white corpuscle in acceptor after bone marrow transplantation.
4. purposes as claimed in claim 3, is characterized in that, described reagent comprises (i) primer or primer pair; (ii) probe; (iii) nucleic acid chip.
5. purposes as claimed in claim 4, is characterized in that, the sequence of described primer pair is as shown in SEQIDNO.:3 and SEQ ID NO.:4.
6. a primer pair of specific amplification STR locus G2S0002, is characterized in that, the sequence of described primer pair is as shown in SEQ ID NO.:3 and SEQ ID NO.:4.
7. a test kit, is characterized in that, described test kit comprises:
(a) for the primer pair of specific amplification STR locus G2S0002;
(b) working instructions.
8. test kit as claimed in claim 7, is characterized in that, described test kit also comprises that (c) specific amplification is selected from the primer pair of one or more locus of lower group:
D3S1358、vWA、D16S539、D5S818、D7S820、CSF1PO、D13S317、TPOX、D8S1179、D21S11、D18S51、TH01、FGA、D2S1338、D19S433、AMELO。
9. an amplification system, is characterized in that, described amplification system comprises:
(a) for the primer pair of specific amplification STR locus G2S0002; With
(b) optional, specific amplification is selected from the primer pair of one or more locus of lower group:
D3S1358、vWA、D16S539、D5S818、D7S820、CSF1PO、D13S317、TPOX、D8S1179、D21S11、D18S51、TH01、FGA、D2S1338、D19S433、AMELO；
And described primer pair (a) and optional primer pair (b) are positioned at same amplification system.
10. amplification system as claimed in claim 9, is characterized in that, described amplification system is composite amplification system, and wherein, described primer pair (b) comprises 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15, or 16 pairs of Auele Specific Primers pair; And/or
Described composite amplification system is fluorescently-labeled composite amplification system, wherein, described fluorescent mark refers to the primer of locus described in one or more fluorescent labeling reagent marks of selecting lower group: FAM, HEX, VIC, NED, PET, TAMRA, ROX, fluorescein, FITC, IRD-700/800, CY3, CY5, CY3.5, CY5.5, TET, TAMRA, JOE, BODIPY TMR, Oregon is green, rhodamine is green, rhodamine is red, texas Red or Ya Jima Huang.
11. 1 kinds of amplification methods, is characterized in that, comprise step:
In claim 9-10, in arbitrary described amplification system, taking sample to be detected as template, utilize polymerase chain reaction to carry out the amplification of locus, thereby obtain locus amplified production.
12. 1 kinds of genetic affinity analytical procedures, is characterized in that, comprise step:
(1) in claim 9-10, in arbitrary described amplification system, taking sample to be detected as template, utilize polymerase chain reaction to carry out the amplification of locus, thereby obtain locus amplified production; With
Wherein, described sample to be detected comprises the sample of nucleic acid S1 from Different Individual respectively, S2 ..., Si, the positive integer that in formula, i is >=2;
(2) to from different IPs acid sample S1, S2 ..., the locus amplified production of Si detects, thereby obtains the detected result corresponding to different IPs acid sample;
(3) detected result step (2) being obtained compares, thereby determines the genetic affinity between Different Individual.
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| CN201310041317.5A CN103966207B (en) | 2013-02-01 | 2013-02-01 | New short nucleotides tandem repetitive sequence site and its application |
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