CN107937571A - A kind of nucleic acid mass spectrum method for paternity test based on information SNP collection and its primer - Google Patents
A kind of nucleic acid mass spectrum method for paternity test based on information SNP collection and its primer Download PDFInfo
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Abstract
The invention discloses a kind of nucleic acid mass spectrum method for paternity test based on information SNP collection and its primer, the information SNP collection used in this method is the data analysis based on 40 crowds, has higher heterozygosis rate>0.4, population genetic index Fst<0.06, and the relatively low frequency of mutation<0.01%, random crowd's matching rate<10‑15It is adapted to individual identity identification and paternity test, the nucleotide sequence of the sense primer of wherein 30 information SNP markers is successively as shown in SEQ ID NO.1~30, the nucleotide sequence of anti-sense primer is successively as shown in SEQ ID NO.31~60, and Single base extension primer nucleotide sequences are successively as shown in SEQ ID NO.61~90.By flight mass spectrum detection information SNP is polymorphic and parent-offspring infers verification experimental verification, qualification result coincide with actual affiliation；Information SNP collection provided by the invention is used for mankind's paternity identification, as a result accurately, preferably by mass spectrometry method, can quickly, the carry out Genotyping of high throughput, simplicity, so as to calculate paternity index and carry out parent-offspring's judgement, have that accuracy is high, the advantages such as cost is low.
Description
Technical field
The present invention relates to using nucleic acid Mass Spectrometer Method information SNP marker carry out mankind's paternity identification detection architecture, specifically
It is related to 30 information SNP marker combinations and primer sequence and method for paternity test.
Background technology
Paternity identification (Parentage Testing) is the detection by being marked to human inheritance, according to genetic development point
Analysis, the identification to genetic connection between individual.The method of modern paternity identification uses DNA analysis, is based primarily upon capillary at present
The PCR-STR composite fluorescence augmentation detections of electrophoretic techniques (Capillary Electrophoresis, CE), it is to apply second
For genetic marker short tandem repeat (STR) typing method, i.e., by carrying out the detection of 15-20 STRs to DNA sample, dividing
Type achievees the purpose that to calculate parental right probability with counting.Str locus seat due to polymorphism is high, contain much information, it is widely distributed and
The advantages that following Mendel's codominant inheritance is considered as the highest genetic marker of versatility in the identification of Forensic DNA.However, with
Extensive use of the str locus seat in forensic identification, its defect also increasingly highlight, such as：The high mutation rate of str locus seat is parent
The result of power identification, which is explained, brings puzzlement；PCR amplification is not easy to realize the DNA typing to degraded；The quantity limitation of STR is unfavorable for multiple
The identification of miscellaneous affiliation.Further, since the sequence length of STR not iso-alleles is similar, can so cause to sentence type error rate
Up to 1%-5%, this will directly affect paternity identification result interpretation.
SNP (Single Nucleotide Polymorphism, single nucleotide polymorphism) is as the third generation after STR
Genetic marker is concerned by people more and more, and SNP typing methods are applied in molecule diagnosis, clinical examination, legal medical expert at present
, genetic disease research, instruct personalized medicine and new drug development etc. many-sided, and gradually substitutes STR, becomes preferred something lost
Mark is passed, because it enriches with widely distributed, quantity, inheritance stability, high throughput, quick and precisely etc. other marks can not compare for detection
The advantage of plan, especially for degraded sample, has more preferable discrimination, therefore become emerging in paternity test field than STR
Popular identification method.
As sequencing technologies develop and investigate crowd size's increase, SNP resolution ratio is also higher and higher in genome.Research
It was found that there is some information SNP (being referred to as Tag SNP) in genome, it can represent other SNP, so, without to base
Because all SNP sites do genetic analysis in group, as long as information SNP site is identified, it is possible to distinguish other in individual
Genotype on site, these sites are referred to as information SNP site or Tag SNP.Research is found：The multiple SNPs mutually adjoined it
Between tend to a stable model genetic to offspring, and multiple SNP sites in certain region of same chromosome
Integrated mode is defined as haplotype.In theory, existence information SNP and substantial amounts of non-information SNP in haplotype, only need to be to information
SNP site is identified, it is possible to knows other common SNPs genotype on haplotype.Therefore need not when doing Genotyping
Parting is carried out to all SNP sites.The hereditary information entrained by 10,000,000 common SNPs in full-length genome can be a small number of
Information SNP is substituted.Therefore choose information SNP site set, more SNP information can be included, solve because SNP quantity is few and
The problem of information content is insufficient, and amplified production length is can be controlled within 200bp, multiplexed PCR amplification easy to implement.Believe in selection
When ceasing SNPs, screening principle is：Higher heterozygosis rate and relatively low crowd variation.Population Genetics index Fst is as a people
The measurement standard of group's allelic variation.Select the final average heterozygosis rates of SNPs>0.45, Fst<0.06, and between two SNP
Average distance more than 37.5Mb, this is also beneficial to the design of later stage nucleic acid Mass Spectrometer Method primer.
Nucleic acid mass spectrometry system, available for quick analysis sample of nucleic acid, its flux disclosure satisfy that more flexible detection limit
Demand, quickly goes out result, has the ability of low cost hundreds of gene mutations of detection.After the system is released, SNP bases are widely used in
Because type analysis and DNA methylation are analyzed.
The content of the invention
In order to overcome the above-mentioned deficiency of the prior art, the purpose of the present invention is for information SNP collection high throughput, contain much information
The advantages of, a set of nucleic acid Mass Spectrometer Method information SNP collection and its multiple PCR primer group available for mankind's paternity identification is established, and
Establish the easy, method for paternity test accurately based on nucleic acid Mass Spectrometer Method information SNP collection.
To reach above-mentioned purpose, technical scheme provides a kind of nucleic acid Mass Spectrometer Method information of mankind's paternity identification
SNP collection, it includes 30 information SNP markers, of the invention 30 information SNP collection are the data analyses based on 40 crowds, tool
There are higher heterozygosis rate ＞ 0.4, population genetic index Fst ＜ 0.06, and relatively low frequency of mutation ＜ 0.01%, random crowd
With rate ＜ 10-15, and the average distance between two SNP is adapted to individual identity identification and paternity test more than 37.5Mb.This hair
30 information SNP markers of bright proposition, position, Fst indexes and its average heterozygosis rate are as shown in table 1.
1 30 information SNP site information of table
| SNP | Chromosome | Chromosomal region band | Position | Fst indexes | Average heterozygosis rate |
| rs560681 | 1 | q23.3 | 157,599,743 | 0.035 | 0.434 |
| rs7520386 | 1 | p36 | 13,900,708 | 0.045 | 0.477 |
| rs12997453 | 2 | q31.3 | 182,238,765 | 0.048 | 0.475 |
| rs6444724 | 3 | q29 | 194,690,082 | 0.045 | 0.468 |
| rs279844 | 4 | p12 | 46,170,583 | 0.03 | 0.485 |
| rs6811238 | 4 | q32.3 | 170,038,345 | 0.031 | 0.485 |
| rs13134862 | 4 | q21.1 | 76,783,075 | 0.054 | 0.471 |
| rs13182883 | 5 | q31 | 136,661,237 | 0.033 | 0.471 |
| rs338882 | 5 | qter | 178,623,331 | 0.056 | 0.454 |
| rs315791 | 5 | q35 | 169,668,498 | 0.058 | 0.449 |
| rs1358856 | 6 | q22 | 123,936,677 | 0.042 | 0.473 |
| rs1336071 | 6 | q16.1 | 94,593,976 | 0.045 | 0.472 |
| rs13218440 | 6 | p24.1 | 12,167,940 | 0.047 | 0.468 |
| rs2272998 | 6 | q24.3 | 148,803,149 | 0.047 | 0.445 |
| rs214955 | 6 | q25 | 152,789,820 | 0.049 | 0.456 |
| rs2503107 | 6 | q22.3 | 127,505,069 | 0.058 | 0.471 |
| rs1019029 | 7 | p22 | 13,667,516 | 0.045 | 0.472 |
| rs10092491 | 8 | p21 | 28,466,991 | 0.039 | 0.456 |
| rs740598 | 10 | q26 | 118,496,889 | 0.04 | 0.463 |
| rs1410059 | 10 | q24.3 | 97,162,585 | 0.054 | 0.467 |
| rs10488710 | 11 | q23.2 | 114,712,386 | 0.025 | 0.441 |
| rs6591147 | 11 | q23 | 105,418,194 | 0.059 | 0.468 |
| rs1058083 | 13 | q32.3 | 98,836,234 | 0.032 | 0.464 |
| rs1821380 | 15 | q13 | 37,100,694 | 0.042 | 0.464 |
| rs7229946 | 18 | q11.1 | 20,992,999 | 0.043 | 0.464 |
| rs9951171 | 18 | p11.3 | 9,739,879 | 0.044 | 0.474 |
| rs985492 | 18 | q11.2 | 27,565,032 | 0.059 | 0.465 |
| rs445251 | 20 | p12.1 | 15,072,933 | 0.041 | 0.463 |
| rs1523537 | 20 | q13.1 | 50,729,569 | 0.042 | 0.472 |
| rs2567608 | 20 | p11.1 | 22,965,082 | 0.044 | 0.475 |
Wherein, in above-mentioned 30 information SNP marker multi-primers group, the nucleotide sequence of sense primer is successively such as SEQ ID
Shown in NO.1~30, the nucleotide sequence of anti-sense primer successively as shown in SEQ ID NO.31~60, Single base extension primer
Nucleotide sequence is successively as shown in SEQ ID NO.61~90.The primer sets sequence of the above-mentioned 30 information SNP markers such as institute of table 2
Show.
The data analysis based on 40 crowds of above-mentioned 30 information SNP markers and obtained by rigorous experiment sieving
, the information SNP collection of final choice has higher heterozygosis rate>0.4th, population genetic index Fst<0.06th, relatively low mutation frequency
Rate<0.01% and random crowd's matching rate<10-15, meet the screening principle of information SNPs, the present invention is also by a whole set of information SNP
Marking the experiment for carrying out parent-offspring's deduction verification in actual colony to demonstrate above-mentioned 30 information SNP markers combination can be accurate
Verification is inferred to mankind's parent child relationship.
The present invention finally provides a kind of method for paternity test based on nucleic acid Mass Spectrometer Method information SNP collection, its feature exists
In comprising the following steps：(1) genome is extracted：Including genomic DNA in extraction anticoagulation DNA and extraction mouth epithelial cells；
(2) multiplex PCR, nucleic acid matter are carried out using the nucleic acid Mass Spectrometer Method information SNP collection of the mankind's paternity identification of claim 1 or 2
Spectrum carries out Genotyping；(3) calculate paternity index and carry out parent-offspring's judgement.
Compared with prior art, the beneficial effects of the present invention are：One kind of the present invention is based on nucleic acid Mass Spectrometer Method information
The method for paternity test of SNP collection, the information SNP collection used in this method is the data analysis based on 40 crowds, is had higher
Heterozygosis rate>0.4, population genetic index Fst<0.06, and the relatively low frequency of mutation<0.01%, random crowd's matching rate<10-15, it is adapted to individual identity identification and paternity test；By flight mass spectrum detection information SNP is polymorphic and parent-offspring infers that experiment is tested
Card, qualification result coincide with actual affiliation；The method for paternity test based on nucleic acid Mass Spectrometer Method information SNP collection of the present invention
High, the advantages such as high throughput and cost are low with accuracy.
Embodiment
Show that example illustrates certain embodiments of the present invention, and should not be construed as the model of the limitation present invention
Enclose.Present disclosure can be improved from material, method and reaction condition at the same time, all these improvement should all
Fall within the spirit and scope of the present invention.No special explanation, the reagent that the embodiment of the present invention uses is commercial goods, this
The database that inventive embodiments use is disclosed online database.
Embodiment 1：The selection of 30 information SNP markers
Selection of the present invention to 30 information SNP markers for paternity identification system is based on essentially from Africa, Europe
The data analysis of 2100 individuals in continent, America and asian population, have passed through three processes, specific as follows：
1) primary dcreening operation goes out heterozygosis rate from demographic data SNPs>0.45, Fst<0.01 SNPs quantity is 2723；
2) on the basis of high heterozygosis rate and low Fst indexes is selected, SNPs of the positional distance less than 1Mb is not selected, while not
The site on X or Y chromosome is selected, obtains the SNPs of 195 primary dcreening operations；
3) heterozygosis rate is screened in the SNPs of 195 primary dcreening operations>0.4th, population genetic index Fst<0.06 and the frequency of mutation<
0.01% SNPs, so as to have selected 30 optimal SNPs as shown in Table 1.
Embodiment 2：Paternity identification is carried out based on nucleic acid Mass Spectrometer Method information SNP collection
1. genome extracts
Using paramagnetic particle method extraction genomic DNA in the present embodiment, sample is saliva, and the saliva for gathering 2ml is stored in saliva
In stabilizer, saliva extracting genome DNA step is as follows：
1) 450 μ l lysates the V, (mixing of saliva and saliva preservation liquid of 450 μ l samples are added in 1.5ml centrifuge tubes
Liquid), 220 μ l isopropanols, 20 μ l Proteinase Ks and 10 μ l nucleic acid settling agents, be vortexed 56 DEG C of water-bath 10min of centrifuge tube after mixing,
Period overturns 6-8 times.
2) centrifuge tube is taken out, 20 μ l magnetic beads is added and is slightly vortexed after mixing, in 56 DEG C of water-baths 20 minutes, during which every 2min
It is vortexed 1 time.
3) take out centrifuge tube and be positioned over magnetic 2min on magnetic frame, supernatant is abandoned in suction.
4) 900 μ l mortifier remover IR are added, be vortexed 30s after mixing, and is placed into magnetic 2min on magnetic frame, and suction is abandoned
Clearly.
5) repeat step (4).
6) 900 μ l rinsing liquid WB are added, are vortexed after mixing 30s, are placed into magnetic 2min on magnetic frame, supernatant is abandoned in suction.
7) repeat step (6).
8) tube cover is opened, dries 3-5min at room temperature, adds 70-120 μ l elution Buffer (advance 56 DEG C of water-baths
10min), it is vortexed and mixes 1min, be then placed into magnetic 2min on magnetic frame, draws DNA and be transferred in new EP pipes.
9) survey concentration using Nanodrop2000 and Qubit3.0 and record, it is ensured that genomic DNA concentration>10ng/ μ l,
A260/A280 is between 1.7~2.0, in -20 DEG C of preservations.Unqualified sample need to extract again.
2.384 hole multiplexed PCR amplification
1) multiplex PCR process is identical with regular-PCR process, according to table 3 by Primer mix, PCR buffer solutions, dNTP
mix、MgCl2, Taq enzyme and water add reaction system, the volume of the extra increase by 38% of all reagents, to prevent in pipetting processes
Loss.
3 multiplexed PCR amplification reaction system of table
2) genomic DNA that 1 μ l concentration is 10ng/ μ l is added to every hole of 384 orifice plates, by the 4 above-mentioned reaction mixtures of μ l
It is added in each hole of 384 orifice plates, slight be vortexed mixes, brief centrifugation.
3) 384 orifice plates sealing plate film is sealed, puts and amplified reaction is carried out in PCR instrument, program is as shown in table 4：
4 multi-PRC reaction loop parameter of table
3. digested using SAP enzymes (shrimp alkaline phosphatase, shrimp alkaline phosphotase)
This step is to digest remaining dNTPs in above-mentioned amplified reaction, prevent influence subsequent reactions, specifically include with
Lower step：
1) according to the preparation SAP enzymic digestions of table 5 reagent, (all reagents have additionally increased by 38% body in 1.5ml centrifuge tubes
Product, prevents the loss in pipetting processes).
5 SAP enzymic digestion reaction systems of table
| Reagent | Volume (1 reaction) | Volume (384 reaction) |
| Water (chromatographically pure) | 1.53μl | 810.9μl |
| SAP buffer solutions (10 ×) | 0.17μl | 90.1μl |
| SAP enzymes (1.7U/ul) | 0.30μl | 159.0μl |
| Cumulative volume | 2.00μl | 1060.0μl |
2) SAP mixed liquors vortex oscillation is mixed for 5 seconds.
3) SAP mixed liquors are centrifuged 10 seconds in 5000rpm, it is every to 384 orifice plates to dispense 2 μ l SAP mixed liquors using point sample instrument
In a hole；
4) 384 orifice plates sealing plate film is sealed, puts and digestion reaction is carried out in PCR instrument, program is as shown in table 6：
6 SAP enzymic digestion reaction cycle parameters of table
| Temperature | Time | Period |
| 37℃ | 40min | 1 |
| 85℃ | 5min | 1 |
| 4℃ | forever | 1 |
4. single base extension
1) SAP reaction plates are put into centrifuge 3000 and leave heart 3min；According to table 7 Suo Shi in 1.5ml centrifuge tubes plus
Enter iPLEX Buffer Plus (10 ×), iPLEX Termination mix, Primer mix, iPLEX enzymes and water reactant
System, the volume that all reagents have added 38% prevent the loss in pipetting processes；
7 single base extension system of table
2) using in each hole of point sample instrument packing 2 μ l reaction mixtures to 384 orifice plates；
3) 384 orifice plates sealing plate film is sealed, puts and single base extension is carried out in PCR instrument, program is as shown in table 8：
8 single base extension loop parameter of table
5. purifying resin and Data Collection
The water dilution of 16 μ l is added into above-mentioned reaction product, the resin being ready for is added and carries out desalting and purifying；Will be pure
To in detection chip, upper machine carries out Mass Spectrometer Method and simultaneously obtains SNP genotyping results sample point sample after change processing.
6. data analysis
SNP typing datas are analyzed, according to 9 formula of table calculate calculate paternity index (paternity index,
PI).When multiple genetic markers are used for paternity test, if the paternity index of each genetic marker is respectively PI1, PI2, PI3 ...
The paternity index of PIn, n genetic markers is multiplied then to add up paternity index (Combinedpaternity index, CPI),
Then：
CPI=PI1 × PI2 × PI3 × ... × PIn (1,2,3, n represent the 1st, 2,3, the PI values of n locus)
9 dyad autosome paternity index calculation formula of table
Note：P, q, r represent the distribution frequency of allele P, Q, R respectively
(1) when being detected the accumulative paternity index of man less than 0.0001, support that it is not child's biology to be detected man
The hypothesis of father.Expert opinion can be expressed as：According to available data and DNA analysis as a result, it is child to exclude to be detected man
Biology father.
(2) when being detected the accumulative paternity index of man more than 10000, support that it is child biology father to be detected man
Hypothesis.Expert opinion can be expressed as：According to available data and DNA analysis as a result, supporting to be detected the biology that man is child
Learn father.
The SNP site number of CPI value=0 and PI=0 more than or equal to 3, can with paternity exclusion, CPI value=0 and
The SNP site number of PI=0 is 1 or 2, not can determine that what can not be excluded for set membership.
Embodiment 3：Application in actual colony
The feasibility and accuracy of paternity identification, this implementation are carried out to verify a whole set of information SNP collection in actual family
Example acquires 20 groups of samples, wherein it is that set membership clearly combines to have 10 groups, remaining 10 groups for unrelated unknown male with it is any
The combinations of pairs of child.It is KY-1~KY-20 that this 20 groups of samples are numbered respectively.Using embodiment 2 obtain data result into
Father and son's affiliation of having gone is inferred, the results are shown in Table 10.
Application of the 10 30 information SNP collection of table in practical colony
| Combination number | Accumulative paternity index (CPI) | Patriarchy is with respect to chance (RCP) | Actual parental right relation |
| KY-1 | 856324110 | 1 | It is one's own |
| KY-2 | 625341255.97 | 1 | It is one's own |
| KY-3 | 965423826.91 | 1 | It is one's own |
| KY-4 | 0 | 0 | It is fictitious |
| KY-5 | 0 | 0 | It is fictitious |
| KY-6 | 3.27E-13 | 3.27E-13 | It is fictitious |
| KY-7 | 7.53684515E-14 | 7.53684515E-14 | It is fictitious |
| KY-8 | 2357891455 | 99.99999901% | It is one's own |
| KY-9 | 65289712234 | 1 | It is one's own |
| KY-10 | 0 | 0 | It is fictitious |
| KY-11 | 0 | 0 | It is fictitious |
| KY-12 | 0 | 0 | It is fictitious |
| KY-13 | 1.91532452E-18 | 1.91532452E-18 | It is fictitious |
| KY-14 | 56422522235.65 | 1 | It is one's own |
| KY-15 | 0 | 0 | It is fictitious |
| KY-16 | 658921355535 | 99.999999902% | It is one's own |
| KY-17 | 489238587464 | 99.999999901% | It is one's own |
| KY-18 | 989835796575 | 1 | It is one's own |
| KY-19 | 0 | 0 | It is fictitious |
| KY-20 | 896181185228 | 1 | It is one's own |
Note：Patriarchy is with respect to chance (RCP)：Since paternity index is real number, it is not easy to find out the chance that you weigh, custom by numeral
Paternity index is converted into a conditional probability, i.e. with respect to chance, and parent child relationship probability, the doubtful father of your table is for patriarchy
Child gives birth to your power of a test size.
As can be seen from Table 10,30 information SNP collection provided by the invention and its multiple PCR primer group are used for mankind's parental right
During identification, its parental right relation is notable, identifies that actual parental right relation fits like a glove.
A kind of method for paternity test based on nucleic acid Mass Spectrometer Method information SNP collection of the present invention, the letter used in this method
It is the data analysis based on 40 crowds to cease SNP collection, has higher heterozygosis rate>0.4, population genetic index Fst<0.06, and
The relatively low frequency of mutation<0.01%, random crowd's matching rate<10-15, it is adapted to individual identity identification and paternity test；By flight
Mass Spectrometer Method information SNP is polymorphic and parent-offspring infers verification experimental verification verification, and qualification result coincide with actual affiliation；The present invention
The method for paternity test based on nucleic acid Mass Spectrometer Method information SNP collection have that accuracy is high, the advantages such as cost is low.
The foregoing is only a preferred embodiment of the present invention, but protection scope of the present invention be not limited thereto,
Any one skilled in the art the invention discloses technical scope in, technique according to the invention scheme and its
Inventive concept is subject to equivalent substitution or change, should be covered by the protection scope of the present invention.
Sequence table
<110>Good training gene biological science and technology（Wuhan）Co., Ltd
<120>A kind of nucleic acid mass spectrum method for paternity test based on information SNP collection and its primer
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<213>Artificial sequence (Artificial Sequence)
<400> 15
acgttggatg ctttattctg gcagcccttg 30
<210> 16
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 16
acgttggatg aaataagtaa ttatatttat 30
<210> 17
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 17
acgttggatg atgatagtgg cttcattttc 30
<210> 18
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 18
acgttggatg attctgtgat agagtggcat 30
<210> 19
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 19
acgttggatg ttcgtaagta tttcaaatag 30
<210> 20
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 20
acgttggatg tcagctgaag gtagacggga 30
<210> 21
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 21
acgttggatg gaggttataa aagactttca 30
<210> 22
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 22
acgttggatg aagatagcct tccacctttt 30
<210> 23
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 23
acgttggatg aggttctgga gttctccatg 30
<210> 24
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 24
acgttggatg tatggaacct ctgctgtctt 30
<210> 25
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 25
acgttggatg aaatgaaaga aggaaacatc 30
<210> 26
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 26
acgttggatg ttgttcctct gggatgcaac 30
<210> 27
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 27
acgttggatg cgaggtgagc ccagaggacc 30
<210> 28
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 28
acgttggatg ccactgcacc tggcctacaa 30
<210> 29
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 29
acgttggatg ttcatttctg catgggtggg 30
<210> 30
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 30
acgttggatg ctgcgccaac cccattctct 30
<210> 31
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 31
acgttggatg agtcaggatg caaactcttg 30
<210> 32
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 32
acgttggatg gaagactctg tcccagccac 30
<210> 33
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 33
acgttggatg accatttaac agctctgatg 30
<210> 34
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 34
acgttggatg tcctactccg tagtaaatga 30
<210> 35
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 35
acgttggatg tcctactccg tagtaaatga 30
<210> 36
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 36
acgttggatg aagtacttct atgaaaatga 30
<210> 37
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 37
acgttggatg aggtcttctt ggtatagctc 30
<210> 38
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 38
acgttggatg cgttactttc ttcctgcctt 30
<210> 39
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 39
acgttggatg catttttctc tccttctgtc 30
<210> 40
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 40
acgttggatg cgcacctctt atttgctcct 30
<210> 41
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 41
acgttggatg aaattatctt ataaactgca 30
<210> 42
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 42
acgttggatg atacctgaaa gcatattaaa 30
<210> 43
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 43
acgttggatg tcaagcagcc ccaacccatc 30
<210> 44
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 44
acgttggatg agcgactcct acgagagaag 30
<210> 45
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 45
acgttggatg gattttcatc aactttatcg 30
<210> 46
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 46
acgttggatg ctaggtctcc acaaccattt 30
<210> 47
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 47
acgttggatg aggataagct cagcctactc 30
<210> 48
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 48
acgttggatg accagtatcc ccgcaaacta 30
<210> 49
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 49
acgttggatg cagctttgca gggccagggt 30
<210> 50
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 50
acgttggatg cttggatatc cttctagctt 30
<210> 51
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 51
acgttggatg actgtattag gagttcccac 30
<210> 52
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 52
acgttggatg ctcctcagcc ctggtctgct 30
<210> 53
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 53
acgttggatg ttgttcttct ccatcccatt 30
<210> 54
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 54
acgttggatg gggcacaatg gagccactga 30
<210> 55
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 55
acgttggatg cagacttgga taaagcagtc 30
<210> 56
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 56
acgttggatg tgtccctgct ttcatgctgg 30
<210> 57
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 57
acgttggatg tcctcctcta aaccaaggga 30
<210> 58
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 58
acgttggatg acatgaacaa attatagtct 30
<210> 59
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 59
acgttggatg tgagacaatg cacagaactg 30
<210> 60
<211> 30
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 60
acgttggatg gcaccttcca ggaggcagca 30
<210> 61
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 61
ctcctgtgac ctgagtaaat 20
<210> 62
<211> 17
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 62
tgtttggtga gctgtat 17
<210> 63
<211> 23
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 63
tgcaagaaag gtaggtaaaa act 23
<210> 64
<211> 26
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 64
aaccaaattg ttgaacactg gttact 26
<210> 65
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 65
gaagtttaga gagttgtgag t 21
<210> 66
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 66
aagccaggtt tgtttaaagt 20
<210> 67
<211> 29
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 67
gtcagatata tcttagatga agcaatagg 29
<210> 68
<211> 16
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 68
ccccaccctg ttcctt 16
<210> 69
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 69
tcaccttctt tcgtgtgcct gtgca 25
<210> 70
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 70
aaaatgagga aactaatgca taggc 25
<210> 71
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 71
ctaagcatcc tgtgtacata t 21
<210> 72
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 72
aagaatgaat ttgaaaaagc atcag 25
<210> 73
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 73
ctgagattca cctctagtcc ctctg 25
<210> 74
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 74
ggtcgtcagt gtggtcagag g 21
<210> 75
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 75
ggtgaacaaa gactgaaaag gtgat 25
<210> 76
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 76
gacaacttga gaaagttagg tatat 25
<210> 77
<211> 28
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 77
ggtcaacaca agatagaagc agactagg 28
<210> 78
<211> 26
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 78
cttccagata gagctaaaac tgaagt 26
<210> 79
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 79
cattgctcgt ctatggttag tctcg 25
<210> 80
<211> 24
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 80
cgggaaacat ctagcatttt tctt 24
<210> 81
<211> 16
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 81
tctgcaaatg tggcag 16
<210> 82
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 82
agattgtcat aactctggac gtatg 25
<210> 83
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 83
tgggttaatt ttgctcagag tatcc 25
<210> 84
<211> 28
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 84
tcttctgtga ttttatattc ttatttat 28
<210> 85
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 85
ccagcaaaca tgtaaagtgt gagag 25
<210> 86
<211> 27
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 86
ggacatgttc acttgtggca gggcaat 27
<210> 87
<211> 25
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 87
agaaatgccc aaaagacttc aggaa 25
<210> 88
<211> 18
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 88
aaaaactgca agtggttg 18
<210> 89
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 89
aacaagatct tgtagggat 19
<210> 90
<211> 17
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 90
tacaacttcc gccgatt 17
Claims (3)
- A kind of 1. nucleic acid Mass Spectrometer Method information SNP collection of mankind's paternity identification, it is characterised in that including 30 information SNP sites, Respectively：rs560681、rs7520386、rs12997453、rs6444724、rs279844、rs6811238、rs13134862、 rs13182883、rs338882、rs315791、rs1358856、rs1336071、rs13218440、rs2272998、 rs214955、rs2503107、rs1019029、rs10092491、rs740598、rs1410059、rs10488710、 rs6591147、rs1058083、rs1821380、rs7229946、rs9951171、rs985492、rs445251、 rs1523537、rs2567608。
- A kind of 2. nucleic acid Mass Spectrometer Method information SNP collection of mankind's paternity identification according to claim 1, it is characterised in that institute The multiple PCR primer group for stating information SNP collection is：Sense primer nucleotide sequence, SEQ ID shown in SEQ ID NO.1~30 The Single base extension prime nucleotide shown in anti-sense primer nucleotide sequence, SEQ ID NO.61~90 shown in NO.31~60 Sequence.
- 3. a kind of nucleic acid mass spectrum method for paternity test based on information SNP collection, it is characterised in that comprise the following steps：(1) extract Genome：Including genomic DNA in extraction anticoagulation DNA and extraction mouth epithelial cells；(2) using described in claim 1 or 2 The nucleic acid Mass Spectrometer Method information SNP collection of mankind's paternity identification carries out multiplex PCR, and nucleic acid mass spectrum carries out Genotyping；(3) parent is calculated Power index simultaneously carries out parent-offspring's judgement.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109628564A (en) * | 2019-02-25 | 2019-04-16 | 北京市理化分析测试中心 | A method of for detecting the primer sets of SNP polymorphism and detecting SNP polymorphism using primer sets |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103173557A (en) * | 2013-04-08 | 2013-06-26 | 上海邃志生物科技有限公司 | Multiple PCR (polymerase chain reaction) primer combination and detection method used for human paternity test |
| CN106536752A (en) * | 2014-03-14 | 2017-03-22 | 凯尔迪克斯公司 | Methods of monitoring immunosuppressive therapies in transplant recipient |
| CN107217095A (en) * | 2017-06-15 | 2017-09-29 | 广东腾飞基因科技股份有限公司 | The mankind's paternity identification multiple PCR primer group and detection method |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103173557A (en) * | 2013-04-08 | 2013-06-26 | 上海邃志生物科技有限公司 | Multiple PCR (polymerase chain reaction) primer combination and detection method used for human paternity test |
| CN106536752A (en) * | 2014-03-14 | 2017-03-22 | 凯尔迪克斯公司 | Methods of monitoring immunosuppressive therapies in transplant recipient |
| CN107217095A (en) * | 2017-06-15 | 2017-09-29 | 广东腾飞基因科技股份有限公司 | The mankind's paternity identification multiple PCR primer group and detection method |
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| SUHUA ZHANG等: "Parallel Analysis of 124 Universal SNPs for Human Identification by Targeted Semiconductor Sequencing", 《SCI REP》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109628564A (en) * | 2019-02-25 | 2019-04-16 | 北京市理化分析测试中心 | A method of for detecting the primer sets of SNP polymorphism and detecting SNP polymorphism using primer sets |
| CN109628564B (en) * | 2019-02-25 | 2022-02-01 | 北京市理化分析测试中心 | Primer set for detecting SNP polymorphism and method for detecting SNP polymorphism by using primer set |
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| CN107937571B (en) | 2021-12-07 |
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