CN107338308A - Multiple PCR primer system, detection method and the application of heredity susceptibility gene of colorectal cancer detection - Google Patents

Multiple PCR primer system, detection method and the application of heredity susceptibility gene of colorectal cancer detection Download PDF

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CN107338308A
CN107338308A CN201710630355.2A CN201710630355A CN107338308A CN 107338308 A CN107338308 A CN 107338308A CN 201710630355 A CN201710630355 A CN 201710630355A CN 107338308 A CN107338308 A CN 107338308A
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dna
homo sapiens
colorectal cancer
heredity
pcr primer
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CN107338308B (en
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许少飞
何广良
唐毅
张纪斌
何铭辉
赖炳权
罗景燕
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Guangzhou Forevergen Biotechnology Co ltd
Guangzhou Forevergen Medical Laboratory Co ltd
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Guangzhou Forevergendx Technology Co ltd
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Abstract

The invention discloses a kind of multiple PCR primer system, detection method and the application of the detection of heredity susceptibility gene of colorectal cancer.The multiple PCR primer system of the heredity susceptibility gene of colorectal cancer detection, including the combination of the first multiple PCR primer and/or the combination of the second multiple PCR primer；Wherein, the combination of the first multiple PCR primer contains 112 pairs of PCR primers pair, and the sequence of 112 pairs of PCR primers pair is respectively such as SEQ ID NO:1~SEQ ID NO:Shown in 224；The combination of first multiple PCR primer contains 113 pairs of PCR primers pair, and the sequence of 113 pairs of PCR primers pair is respectively such as SEQ ID NO:225~SEQ ID NO:Shown in 450.The multiple PCR primer system of the present invention can detect 11 related tumor susceptibility genes of colorectal cancer simultaneously, interfering property is small between each primer, high specificity, expanding effect is good, can improve detection efficiency, reduces testing cost, and high throughput sequencing technologies can be coordinated, high-flux sequence is carried out, to colorectal cancer Susceptible population, the crowd especially with colorectal cancer household heredity factors has certain auxiliary diagnosis and suggesting effect.

Description

The multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection, detection side Method and application
Technical field
The present invention relates to molecular Biological Detection field, is detected more particularly, to a kind of heredity susceptibility gene of colorectal cancer Multiple PCR primer system, detection method and application.
Background technology
Colorectal cancer (Colorectal Cancer, CRC) is the general name of colon cancer and the carcinoma of the rectum, refers to colorectal mucosa epithelium The canceration occurred under a variety of carcinogenic factor effects such as environment or heredity, is the third-largest common cancer in the whole world, it is dead Rate is higher.Research finds that 35% CRC has family's neurological susceptibility, and most common of which is hereditary nonpolyposis colorectal cancer (hereditary nonpolyposis colorectal cancer, HNPCC), in addition, familial adenomatosis polyposis (familial adenomatous polyposis, FAP) and paramnesia Jeghers syndrome (hamartomatous polyposis Syndromes, HPS) it is also relatively common.Therefore, carry out the analysis to the tumor susceptibility gene of heredity colorectal cancer and study to losing The pathogenesis of transmissibility colorectal cancer and auxiliary early diagnosis have important application value.
The content of the invention
Based on this, it is necessary to provide a kind of multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection, inspection Survey methods and applications.
The technical scheme that the present invention solves above-mentioned technical problem is as follows.
A kind of multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection, including the first multiple PCR primer Combination and/or the combination of the second multiple PCR primer；Wherein, the first multiple PCR primer combination contains 112 pairs of PCR primers pair, And the sequence of 112 pairs of PCR primers pair is respectively such as SEQ ID NO:1~SEQ ID NO:Shown in 224；First multiplex PCR Primer combination contains 113 pairs of PCR primers pair, and the sequence of 113 pairs of PCR primers pair is respectively such as SEQ ID NO:225~SEQ ID NO:Shown in 450.
In one of the embodiments, each pair of primer equimolar amounts mixing in the first multiple PCR primer combination.
In one of the embodiments, each pair of primer equimolar amounts mixing in the second multiple PCR primer combination.
The multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection described in any of the above-described embodiment can answer In the detection reagent for preparing the detection straight colon cancer of heredity.
A kind of heredity susceptibility gene of colorectal cancer detection kit, contains the heredity described in any of the above-described embodiment The multiple PCR primer system of susceptibility gene of colorectal cancer detection.
In one of the embodiments, the heredity susceptibility gene of colorectal cancer detection is also carried with kit including DNA Take reagent, PCR amplifing reagents, and/or DNA purified reagents.
A kind of detection method of heredity susceptibility gene of colorectal cancer, comprises the following steps：
Use the multiple PCR primer system of the heredity susceptibility gene of colorectal cancer detection described in any of the above-described embodiment System, multiplexed PCR amplification is carried out by template of the genomic DNA of acquisition；
Sequencing library is built to the product of multiplexed PCR amplification；
High-flux sequence is carried out to the sequencing library of structure, and sequencing result is compared and arrives human genome reference sequences On.
In one of the embodiments, the structure sequencing library includes successively：End is carried out to the product of multiplexed PCR amplification End is repaired, end adds A, connect sequence measuring joints, amplified library, the library after amplification is purified and fragment is screened.
In one of the embodiments, the human genome reference sequences are hg19.
In one of the embodiments, the detection method of the heredity susceptibility gene of colorectal cancer also includes tying to comparing Fruit carries out the step of Mutation Screening analysis and mutation progress functional annotation to screening.
The present invention has found that hereditary nonpolyposis colorectal cancer, familial adenomatosis polyposis and paramnesia cease by research It is relevant that there is meat syndrome etc. the cancer of family's neurological susceptibility to be mutated with the family of 11 genes, wherein, hereditary nonpolyposis The influence gene of colorectal cancer includes MSH2, MLH1, PMS2, MSH6, EPCAM, and wherein MLH1 occupies the 50% of harmful family mutation, MSH2, which accounts for 40%, MSH6 and accounts for 7%~10%, PMS2, to be accounted for<5%, EPCAM account for 1%, and familial adenomatosis polyposis includes classic FAP and astenic form FAP, it, which influences gene, includes APC, MUTYH, and paramnesia Jeghers syndrome includes juvenile polyposis, cowden Syndrome and Peutz Jeghers syndromes, it, which influences gene, includes BMPR1A, SMAD4, PTEN, STK11.
The multiple PCR primer system of the heredity susceptibility gene of colorectal cancer detection of the present invention can detect Colon and rectum simultaneously 11 related tumor susceptibility genes of cancer, interfering property is small between each primer, and high specificity, expanding effect is good, and primer skewed popularity expands The degree of increasing is low, can improve detection efficiency, reduces testing cost, and can coordinate high throughput sequencing technologies, has detection flux High, accuracy and high sensitivity, it is reproducible the features such as, to colorectal cancer Susceptible population, especially with colorectal cancer family The crowd of Genetic history has certain auxiliary diagnosis and suggesting effect.
Brief description of the drawings
Fig. 1 is the result of 293T-1 libraries 2100 in specific embodiment；
Fig. 2 is the result of 293T-2 libraries 2100 in specific embodiment.
Embodiment
For the ease of understanding the present invention, the present invention is described more fully below with reference to relevant drawings.In accompanying drawing Give presently preferred embodiments of the present invention.But the present invention can realize in many different forms, however it is not limited to this paper institutes The embodiment of description.On the contrary, the purpose for providing these embodiments is to make the understanding to the disclosure more thorough Comprehensively.
Unless otherwise defined, all of technologies and scientific terms used here by the article is with belonging to technical field of the invention The implication that technical staff is generally understood that is identical.Term used in the description of the invention herein is intended merely to description tool The purpose of the embodiment of body, it is not intended that in the limitation present invention.Term as used herein "and/or" includes one or more phases The arbitrary and all combination of the Listed Items of pass.
Embodiment 1
First, design of primers
The target area that the present embodiment design of primers is directed to is the encoded exon of related 11 tumor susceptibility genes of colorectal cancer With the base of exon-intron boundaries 10.Because the most long codes extron of portion gene is up to thousands of, single pair primer can not expand The region of the length, and sequencing reading length does not reach the length, so needing to be divided the excessive extron of length, assigns to more Carried out in individual PCR reaction systems.
The heredity susceptibility gene of colorectal cancer detection multiple PCR primer Survey design of the present embodiment design is as shown in table 1.
The heredity susceptibility gene of colorectal cancer of table 1 detects multiple PCR primer Survey design
2nd, experiment material
The cell line 293T that the present embodiment is related to derives from Guangzhou Sai Ku Bioisystech Co., Ltd, and is answered on request Soviet Union, culture and passage.
All primer purity reach electrophoresis level (PAGE), without miscellaneous band.
Main agents are such as：KOD muti＆Epi and 2 × PCR Buffer for KOD-muti＆Epi (TOYOBO, article No. KME-101), VAHTS Nano DNA Library Prep Kit for Illumina (promise is only praised, article No. ND601-02), VAHTS DNA Adapters set1/set2for Illumina (promise is only praised, article No. N801-01), VAHTS DNA Clean Beads (promise is only praised, article No. N411-02), HiPure Blood＆Tissue DNA Kit (Magen, article No. D3018-03) etc..
Key instrument is such as：The sequenators of Illumina Nextseq 500, Agilent 2100bioanalyzer, Qubit are glimmering Light quantitative instrument, PCR instrument, supercentrifuge, water-bath, whirlpool concussion instrument, refrigerator, pipettor etc..
3rd, multiplexed PCR amplification, library construction and NGS sequencings
225 pairs of primers are mixed into 2 Mix by design, it is rich to carry out target sequence using 293T cell lines as standard items Collection and library construction, if 2 repetitions, respectively 293T-1 and 293T-2.
(1) multiplexed PCR amplification of .DNA templates
1. primer adds sterilizing purified water dissolving, final concentration of 400nM.SEQ ID NO:1~SEQ ID NO:Shown in 224 The material such as primer amount mixing, turn into primer Mix 1, SEQ ID NO:225~SEQ ID NO:The things such as the primer shown in 450 The amount mixing of matter, turns into primer Mix 2.Primer Mix 1/2 final concentration is 50nM.
2. sample DNA extracts.
Use HiPure Blood＆Tissue DNA Kit, the DNA of extracting 293T cells.
3. same sample to be tested is divided into the pipe of A, B two to be reacted.
Amplification system see the table below 2.
Table 2
Reagent Volume (μ l)
2×PCR Buffer for KOD-Multi&Epi 10
KOD-Multi&Epi(1.0U/μl) 0.4
Primer Mix1/2 (50 μM) 7.6
DNA sample (5ng/ μ l) 2
Cumulative volume 20
PCR amplification programs see the table below 3.
Table 3
4. after the amount mixing of the materials such as the pcr amplification product of same sample, (VAHTS DNA Clean are come from using magnetic bead Beads (promise is only praised)) purified.
(2) uses VAHTS Nano DNA Library Prep Kit for Illumina (promise is only praised) kit, Carry out library construction
1. end is repaired：After the mixing of pcr amplification product after purification 20 μ l End Prep Mix and 30 μ l, 30 DEG C incubate Educate 30min.
2. repair product to end using magnetic bead to purify.
3. end adds A：It is equal that product mixing is repaired in 12.5 μ l dA-Tailing Mix and 17.5 μ l end after purification After even, 37 DEG C of incubations 30min, 70 DEG C of incubations 5min, 4 DEG C of incubation 5min.
4. sequence measuring joints connect：2.5 μ l Ligation Mix and 2.5 μ l DNA are added in product into step 3 Adapter X (come from VAHTS DNA Adapters set1/set2for Illumina (promise is only praised)), after being well mixed, 30 DEG C be incubated 10min.
5. purified using magnetic bead butt joint connection product.
6. amplified library：Using connection product after purification as DNA profiling, enter performing PCR amplification.
Amplification system see the table below 4.
Table 4
DNA profiling 20μl
PCR Primer Mix 5μl
Amplification Mix2 25μl
Cumulative volume 50μl
Amplification program see the table below 5.
Table 5
7. the library after amplification is purified using magnetic bead and fragment is screened.
(3) quality inspection of libraries and sequencing
1. library concentration is quantified using Qubit fluorescent quantitation instruments.
2. quality inspection is carried out to Library Quality using Agilent 2100bioanalyzer.
3. sample library is sequenced using the sequenators of Nextseq 500.
4th, experimental result and data analysis
Sequencing data analysis is compared using Bwa softwares with human genome reference sequences hg19, soft with GATK instruments Part carries out Mutation Screening analysis (examination condition to comparison result：1st, compare quality and be more than 20；2nd, depth is sequenced to be more than 100), uses Mutation of the ANNOVAR softwares to screening carries out functional annotation.
Result such as Fig. 1 of the analysis of 293T-1 and 293T-2 high-throughput sequencing libraries 2100,2.It can be seen from Fig. 1 and Fig. 2 The sequencing library of high quality is finally generated, and the distribution of lengths in library is consistent with respective expected design.
Above-mentioned library machine on Illumina Nextseq 500 are carried out is sequenced, and sequencing pattern is PE150.For number is sequenced According to by analysis of biological information, one side statistics index is used to evaluate sequence enrichment effect, on the other hand carries out monokaryon glycosides Sour polymorphism (SNP) and short insertion and deletion (InDel) abrupt climatic change, assess the feasible of heredity susceptibility gene of colorectal cancer detection Property.
The sequencing data of table 6 counts
As shown in table 6, target area size is about 32Kb, and two library 293T-1 and 293T-2 sequencings of the present embodiment are produced More than 3,100,000 read sequences are given birth to, sequencing data amount is respectively 470Mb and 660Mb.Short sequence in sequencing data more than 95% Row can compare genome, target area has been arrived in more than 97% comparison in the data of genome in comparisons, it was demonstrated that multiplex PCR The high degree of specificity of sequence enrichment.In addition, the present embodiment has reached 100% coverage to the concentration effect of target area sequence, And target area average sequencing depth is up to more than 10,000 times.The present embodiment need to only be carried out in 2 PCR reaction systems, on the one hand be subtracted Laboratory operating procedures are lacked, while sample initial amount are also reduced to 20ng (as shown in table 1).
The purpose of design of the multiple PCR primer system of the heredity susceptibility gene of colorectal cancer detection of the present embodiment be for Genetic test is carried out to heredity colorectal cancer, that is, analyzed on heredity susceptibility gene of colorectal cancer with the presence or absence of functional Mutation, including SNP (SNP) and short insertion and deletion (InDel).Sequencing data mutation analysis 293T-1, 293T-2, in detect 15 SNP, be not detected by InDel, the uniformity 100% of two results.By to this 15 SNP Further analysis, its function classification belongs to normal polymorphism.
To sum up, the multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection of the invention be it is a kind of efficiently, PCR primer system is used in reliable high-flux sequence amplification, and it can detect 11 related tumor susceptibility genes of colorectal cancer simultaneously, improve Detection efficiency, and reduce testing cost, also there is to detect flux high, accuracy and high sensitivity, reproducible, to tying Carcinoma of the rectum Susceptible population, the crowd especially with colorectal cancer household heredity factors have certain auxiliary diagnosis and suggesting effect.
Each technical characteristic of embodiment described above can be combined arbitrarily, to make description succinct, not to above-mentioned reality Apply all possible combination of each technical characteristic in example to be all described, as long as however, the combination of these technical characteristics is not deposited In contradiction, the scope that this specification is recorded all is considered to be.
Embodiment described above only expresses the several embodiments of the present invention, and its description is more specific and detailed, but simultaneously Can not therefore it be construed as limiting the scope of the patent.It should be pointed out that come for one of ordinary skill in the art Say, without departing from the inventive concept of the premise, various modifications and improvements can be made, these belong to the protection of the present invention Scope.Therefore, the protection domain of patent of the present invention should be determined by the appended claims.
SEQUENCE LISTING
<110>Guangzhou promise health Science and Technology Ltd. forever, Guangzhou Yong Nuo bio tech ltd
<120>Multiple PCR primer system, detection method and the application of heredity susceptibility gene of colorectal cancer detection
<130> 2017
<160> 450
<170> PatentIn version 3.3
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<212> DNA
<213> Homo sapiens
<400> 54
aaaccatcag caacttcttc catgat 26
<210> 55
<211> 24
<212> DNA
<213> Homo sapiens
<400> 55
agcccttctt ggtagatttc aact 24
<210> 56
<211> 26
<212> DNA
<213> Homo sapiens
<400> 56
aaacagcact acttatcaaa gcctga 26
<210> 57
<211> 24
<212> DNA
<213> Homo sapiens
<400> 57
actgtgtttg gaaaatgatc acct 24
<210> 58
<211> 26
<212> DNA
<213> Homo sapiens
<400> 58
agtgcgtgct ctaaaaacat tcatat 26
<210> 59
<211> 30
<212> DNA
<213> Homo sapiens
<400> 59
gaaacttaag gtcagttagt tataattcca 30
<210> 60
<211> 22
<212> DNA
<213> Homo sapiens
<400> 60
aagtcgtagc ccttaagtga gc 22
<210> 61
<211> 23
<212> DNA
<213> Homo sapiens
<400> 61
cagacaatgt catcacagga gga 23
<210> 62
<211> 28
<212> DNA
<213> Homo sapiens
<400> 62
acaaacaaag cttcaacaat ttactctc 28
<210> 63
<211> 22
<212> DNA
<213> Homo sapiens
<400> 63
accttatctc caccagcaaa ct 22
<210> 64
<211> 23
<212> DNA
<213> Homo sapiens
<400> 64
ggtgtttcct gtgagtggat ttc 23
<210> 65
<211> 22
<212> DNA
<213> Homo sapiens
<400> 65
atttcaaagg ccccagagaa gt 22
<210> 66
<211> 21
<212> DNA
<213> Homo sapiens
<400> 66
catacctggg gttgctggaa g 21
<210> 67
<211> 25
<212> DNA
<213> Homo sapiens
<400> 67
catgactgct ttctccattt ccaaa 25
<210> 68
<211> 26
<212> DNA
<213> Homo sapiens
<400> 68
tggagagcta ctattttcag aaacga 26
<210> 69
<211> 22
<212> DNA
<213> Homo sapiens
<400> 69
cacatgcatg taccgaaatg ct 22
<210> 70
<211> 27
<212> DNA
<213> Homo sapiens
<400> 70
caacactttg tatcggaata cagagaa 27
<210> 71
<211> 26
<212> DNA
<213> Homo sapiens
<400> 71
acctcaagtt tacaagaggg aatact 26
<210> 72
<211> 26
<212> DNA
<213> Homo sapiens
<400> 72
ggagtacaca aggcaatgtt tactat 26
<210> 73
<211> 35
<212> DNA
<213> Homo sapiens
<400> 73
tgtctttaat aaaatttaat caatattaac agagc 35
<210> 74
<211> 26
<212> DNA
<213> Homo sapiens
<400> 74
aaccatcttg cttcatactt ttctga 26
<210> 75
<211> 23
<212> DNA
<213> Homo sapiens
<400> 75
ctctttactg ccccgggaat ttc 23
<210> 76
<211> 31
<212> DNA
<213> Homo sapiens
<400> 76
aagacagcat attaactaaa catgttttct g 31
<210> 77
<211> 29
<212> DNA
<213> Homo sapiens
<400> 77
ctcatacctg agctatctta agaaataca 29
<210> 78
<211> 27
<212> DNA
<213> Homo sapiens
<400> 78
aagccaaaca tgaaattcat attatag 27
<210> 79
<211> 32
<212> DNA
<213> Homo sapiens
<400> 79
aactttcatg attatataaa acattgctta ca 32
<210> 80
<211> 24
<212> DNA
<213> Homo sapiens
<400> 80
tgagatgcct tgggacttaa attg 24
<210> 81
<211> 22
<212> DNA
<213> Homo sapiens
<400> 81
cctgagaggt atgaatggct ga 22
<210> 82
<211> 27
<212> DNA
<213> Homo sapiens
<400> 82
tggtgtatct agttctccat cattatc 27
<210> 83
<211> 26
<212> DNA
<213> Homo sapiens
<400> 83
tggtaggctt atcatcttca tagtca 26
<210> 84
<211> 27
<212> DNA
<213> Homo sapiens
<400> 84
ggagtatctt ctacacaata agtctgt 27
<210> 85
<211> 26
<212> DNA
<213> Homo sapiens
<400> 85
gaagtacatc tgctaaacat gagtgg 26
<210> 86
<211> 22
<212> DNA
<213> Homo sapiens
<400> 86
cactcaggct ggatgaacaa ga 22
<210> 87
<211> 26
<212> DNA
<213> Homo sapiens
<400> 87
cctgttttgt gatggtagaa gtttgt 26
<210> 88
<211> 22
<212> DNA
<213> Homo sapiens
<400> 88
cttgtgactt ttccctttgg gc 22
<210> 89
<211> 26
<212> DNA
<213> Homo sapiens
<400> 89
tcatttcgtg aaaaacagta aggagt 26
<210> 90
<211> 24
<212> DNA
<213> Homo sapiens
<400> 90
tgttttcttg gtcaatgtca ctga 24
<210> 91
<211> 26
<212> DNA
<213> Homo sapiens
<400> 91
ccaatcaaaa ttttctgaat cagggg 26
<210> 92
<211> 22
<212> DNA
<213> Homo sapiens
<400> 92
gcttgaaggg gctgtttcat tt 22
<210> 93
<211> 26
<212> DNA
<213> Homo sapiens
<400> 93
ggatgatgtc cttggaagtt gagata 26
<210> 94
<211> 25
<212> DNA
<213> Homo sapiens
<400> 94
gattcaaatg aagcagattc ctcca 25
<210> 95
<211> 26
<212> DNA
<213> Homo sapiens
<400> 95
tgctttttca ctggattctg atgaag 26
<210> 96
<211> 29
<212> DNA
<213> Homo sapiens
<400> 96
tgcctgatta tcttttgaat ctttaatgt 29
<210> 97
<211> 22
<212> DNA
<213> Homo sapiens
<400> 97
gctgtcagtt ttggaatctc gc 22
<210> 98
<211> 26
<212> DNA
<213> Homo sapiens
<400> 98
ccagccttat tttagagaag aatgct 26
<210> 99
<211> 26
<212> DNA
<213> Homo sapiens
<400> 99
tgttgttttc atttcatttc tgctgt 26
<210> 100
<211> 22
<212> DNA
<213> Homo sapiens
<400> 100
cctctcaggt tgatgtagct gt 22
<210> 101
<211> 24
<212> DNA
<213> Homo sapiens
<400> 101
aaagaggcag tgagttccag tcac 24
<210> 102
<211> 24
<212> DNA
<213> Homo sapiens
<400> 102
ctctcaccat ttcaggatag tccc 24
<210> 103
<211> 26
<212> DNA
<213> Homo sapiens
<400> 103
ttgctacaag aggaaaagct tttgtt 26
<210> 104
<211> 22
<212> DNA
<213> Homo sapiens
<400> 104
gaatgttgtt agaatgcggg gg 22
<210> 105
<211> 26
<212> DNA
<213> Homo sapiens
<400> 105
agtccactct gtctttatta ggaagt 26
<210> 106
<211> 25
<212> DNA
<213> Homo sapiens
<400> 106
atcatgggtc tctcactgaa cacac 25
<210> 107
<211> 22
<212> DNA
<213> Homo sapiens
<400> 107
actgatagca tgggtccgtt tt 22
<210> 108
<211> 20
<212> DNA
<213> Homo sapiens
<400> 108
cccgggtccc tggatgtgcc 20
<210> 109
<211> 20
<212> DNA
<213> Homo sapiens
<400> 109
cctcttggtc ccctccctat 20
<210> 110
<211> 18
<212> DNA
<213> Homo sapiens
<400> 110
cgctaccgat ctccgcag 18
<210> 111
<211> 22
<212> DNA
<213> Homo sapiens
<400> 111
atatcaggac aagcagtgtg gc 22
<210> 112
<211> 18
<212> DNA
<213> Homo sapiens
<400> 112
aacgggtgca gtgcctgt 18
<210> 113
<211> 20
<212> DNA
<213> Homo sapiens
<400> 113
gccaatggga gttcaggagg 20
<210> 114
<211> 19
<212> DNA
<213> Homo sapiens
<400> 114
gacatcctgg ccgagtcag 19
<210> 115
<211> 22
<212> DNA
<213> Homo sapiens
<400> 115
ttaagggcag aacaccggtt ta 22
<210> 116
<211> 22
<212> DNA
<213> Homo sapiens
<400> 116
aagagagaat ggagggaatc gg 22
<210> 117
<211> 22
<212> DNA
<213> Homo sapiens
<400> 117
gtgactctgc cctatgacac tc 22
<210> 118
<211> 19
<212> DNA
<213> Homo sapiens
<400> 118
gtgatctcac agcccaccc 19
<210> 119
<211> 21
<212> DNA
<213> Homo sapiens
<400> 119
atagagggac tgacgggtga t 21
<210> 120
<211> 20
<212> DNA
<213> Homo sapiens
<400> 120
caggtcagca gtgtcctcat 20
<210> 121
<211> 20
<212> DNA
<213> Homo sapiens
<400> 121
ttgggggaag ccctaagtgg 20
<210> 122
<211> 22
<212> DNA
<213> Homo sapiens
<400> 122
tttgaaggct acctctggga ag 22
<210> 123
<211> 27
<212> DNA
<213> Homo sapiens
<400> 123
tctattctcc taactcttaa gaagggc 27
<210> 124
<211> 26
<212> DNA
<213> Homo sapiens
<400> 124
gttcaattct aggtagcctg gttact 26
<210> 125
<211> 27
<212> DNA
<213> Homo sapiens
<400> 125
tctaaatcaa cacaccattc atgtcta 27
<210> 126
<211> 26
<212> DNA
<213> Homo sapiens
<400> 126
gaggatgttt ttgctcttta ggtctc 26
<210> 127
<211> 32
<212> DNA
<213> Homo sapiens
<400> 127
gctgttattt ttaaaatgat gagtaaatca ac 32
<210> 128
<211> 26
<212> DNA
<213> Homo sapiens
<400> 128
atccttattc cactctaatt ccaccc 26
<210> 129
<211> 22
<212> DNA
<213> Homo sapiens
<400> 129
atccgtctac tcccacgttc ta 22
<210> 130
<211> 26
<212> DNA
<213> Homo sapiens
<400> 130
ctctacctca ctctaacaag cagata 26
<210> 131
<211> 22
<212> DNA
<213> Homo sapiens
<400> 131
ttacaccagt tcgtcccttt cc 22
<210> 132
<211> 27
<212> DNA
<213> Homo sapiens
<400> 132
ggttaagaaa actgttccaa tacatgg 27
<210> 133
<211> 26
<212> DNA
<213> Homo sapiens
<400> 133
ggctttgtct ttatttgctt tgtcaa 26
<210> 134
<211> 29
<212> DNA
<213> Homo sapiens
<400> 134
tcaagtttat tttcatggtg ttttatccc 29
<210> 135
<211> 23
<212> DNA
<213> Homo sapiens
<400> 135
gcttgaaagg aaacgtagca agt 23
<210> 136
<211> 26
<212> DNA
<213> Homo sapiens
<400> 136
gctcacacaa actaattcta ggtcaa 26
<210> 137
<211> 24
<212> DNA
<213> Homo sapiens
<400> 137
ggaaaactca acttgctgac taca 24
<210> 138
<211> 24
<212> DNA
<213> Homo sapiens
<400> 138
aacagaaaac aaagccctac caaa 24
<210> 139
<211> 26
<212> DNA
<213> Homo sapiens
<400> 139
tggagtgctt acaaattctg agtttt 26
<210> 140
<211> 26
<212> DNA
<213> Homo sapiens
<400> 140
ccttccaccc agatttcaat tctttt 26
<210> 141
<211> 23
<212> DNA
<213> Homo sapiens
<400> 141
accatcctga taaggttaag ggc 23
<210> 142
<211> 22
<212> DNA
<213> Homo sapiens
<400> 142
aggaagacag aaccatcagc ac 22
<210> 143
<211> 19
<212> DNA
<213> Homo sapiens
<400> 143
tcgtgtgcca gtctccttc 19
<210> 144
<211> 22
<212> DNA
<213> Homo sapiens
<400> 144
actctttcca actcaaggca ca 22
<210> 145
<211> 26
<212> DNA
<213> Homo sapiens
<400> 145
tgaatctgaa cagcatgaaa acaagt 26
<210> 146
<211> 22
<212> DNA
<213> Homo sapiens
<400> 146
tactgtctca ccaactgtct gc 22
<210> 147
<211> 26
<212> DNA
<213> Homo sapiens
<400> 147
cccagccact attacttttt cttgaa 26
<210> 148
<211> 26
<212> DNA
<213> Homo sapiens
<400> 148
aaactcatga ccttcaaaga tgtctt 26
<210> 149
<211> 20
<212> DNA
<213> Homo sapiens
<400> 149
tctctgaggc gggaaaggag 20
<210> 150
<211> 26
<212> DNA
<213> Homo sapiens
<400> 150
attatcaggg aattcacaca gtccta 26
<210> 151
<211> 34
<212> DNA
<213> Homo sapiens
<400> 151
taatatcatt gatacacagt ttaggttttg agat 34
<210> 152
<211> 28
<212> DNA
<213> Homo sapiens
<400> 152
ttagctcctt tataagcttc ttcagtat 28
<210> 153
<211> 22
<212> DNA
<213> Homo sapiens
<400> 153
atcatgtggg taactgcagg tt 22
<210> 154
<211> 22
<212> DNA
<213> Homo sapiens
<400> 154
ctaagagtga gtcaccacca cc 22
<210> 155
<211> 26
<212> DNA
<213> Homo sapiens
<400> 155
tttcttaaag tggcctttgc ttttta 26
<210> 156
<211> 26
<212> DNA
<213> Homo sapiens
<400> 156
agaacattca catcatgtta gagcat 26
<210> 157
<211> 22
<212> DNA
<213> Homo sapiens
<400> 157
gaacttcaac acaagcatgc ct 22
<210> 158
<211> 27
<212> DNA
<213> Homo sapiens
<400> 158
agacatacat ttctatcttc aagggac 27
<210> 159
<211> 25
<212> DNA
<213> Homo sapiens
<400> 159
agggtagtaa gtttcccatt accaa 25
<210> 160
<211> 30
<212> DNA
<213> Homo sapiens
<400> 160
acagacaata gcttatcaat attaccttca 30
<210> 161
<211> 22
<212> DNA
<213> Homo sapiens
<400> 161
tgcctgtctg tctgtttctc tc 22
<210> 162
<211> 25
<212> DNA
<213> Homo sapiens
<400> 162
tcccccatca ccctaacata aataa 25
<210> 163
<211> 26
<212> DNA
<213> Homo sapiens
<400> 163
gctctcaaag tattcttggt ttctga 26
<210> 164
<211> 22
<212> DNA
<213> Homo sapiens
<400> 164
gcccagttgc ctttcatgaa ta 22
<210> 165
<211> 22
<212> DNA
<213> Homo sapiens
<400> 165
ggcgatcatc tgaaaactga cc 22
<210> 166
<211> 25
<212> DNA
<213> Homo sapiens
<400> 166
agctcctgat caataaggca ttttt 25
<210> 167
<211> 22
<212> DNA
<213> Homo sapiens
<400> 167
tagcctctca agatctggaa gc 22
<210> 168
<211> 24
<212> DNA
<213> Homo sapiens
<400> 168
gtcataatca gagtcaaagc ctgc 24
<210> 169
<211> 22
<212> DNA
<213> Homo sapiens
<400> 169
caaagtctta cccaacactg cg 22
<210> 170
<211> 21
<212> DNA
<213> Homo sapiens
<400> 170
gctcctcttc ctcacagcct a 21
<210> 171
<211> 26
<212> DNA
<213> Homo sapiens
<400> 171
aatgactgaa tgagaactta agtggg 26
<210> 172
<211> 23
<212> DNA
<213> Homo sapiens
<400> 172
attagcgata catgtgctag caa 23
<210> 173
<211> 24
<212> DNA
<213> Homo sapiens
<400> 173
ttaccacctt tgtcagaagt caac 24
<210> 174
<211> 22
<212> DNA
<213> Homo sapiens
<400> 174
ttccatgaag cgcacaaaca tc 22
<210> 175
<211> 22
<212> DNA
<213> Homo sapiens
<400> 175
acactggtgt tgagacagga tt 22
<210> 176
<211> 22
<212> DNA
<213> Homo sapiens
<400> 176
caaatctcag agacccactc cc 22
<210> 177
<211> 27
<212> DNA
<213> Homo sapiens
<400> 177
tgaaaggttc caaaataatg tgatgga 27
<210> 178
<211> 24
<212> DNA
<213> Homo sapiens
<400> 178
ggagagcctg atagaacatc tgtt 24
<210> 179
<211> 20
<212> DNA
<213> Homo sapiens
<400> 179
ctctgtgaca atggcctggg 20
<210> 180
<211> 21
<212> DNA
<213> Homo sapiens
<400> 180
ggcaagtcag gcagagagaa g 21
<210> 181
<211> 22
<212> DNA
<213> Homo sapiens
<400> 181
tctccttagc ttttgtgcct gt 22
<210> 182
<211> 27
<212> DNA
<213> Homo sapiens
<400> 182
ccggctggaa attttatttg aagaata 27
<210> 183
<211> 22
<212> DNA
<213> Homo sapiens
<400> 183
atgaggtcct gtcctagtcc tg 22
<210> 184
<211> 19
<212> DNA
<213> Homo sapiens
<400> 184
gggaggagac gtcaatgcc 19
<210> 185
<211> 26
<212> DNA
<213> Homo sapiens
<400> 185
tggatctaca cacctaaaga tgacaa 26
<210> 186
<211> 30
<212> DNA
<213> Homo sapiens
<400> 186
cagttatcct gaattttaat ggattaccta 30
<210> 187
<211> 30
<212> DNA
<213> Homo sapiens
<400> 187
tggtgtcaca ttaaattata catgttttct 30
<210> 188
<211> 26
<212> DNA
<213> Homo sapiens
<400> 188
tagcaaagta gtcatggcat tagtga 26
<210> 189
<211> 25
<212> DNA
<213> Homo sapiens
<400> 189
catacttgga tttttgtcct ggtcc 25
<210> 190
<211> 27
<212> DNA
<213> Homo sapiens
<400> 190
tctaccatga tttaaaaatc caccagt 27
<210> 191
<211> 22
<212> DNA
<213> Homo sapiens
<400> 191
agattgcaca actgccctct aa 22
<210> 192
<211> 26
<212> DNA
<213> Homo sapiens
<400> 192
cctccacttt caataatggc taaagt 26
<210> 193
<211> 25
<212> DNA
<213> Homo sapiens
<400> 193
tcccatagca atcattttgt gcttt 25
<210> 194
<211> 21
<212> DNA
<213> Homo sapiens
<400> 194
tcttgatgaa gaggagctgg g 21
<210> 195
<211> 28
<212> DNA
<213> Homo sapiens
<400> 195
ttgaagatct cttgtattct aatttggc 28
<210> 196
<211> 24
<212> DNA
<213> Homo sapiens
<400> 196
aggtgtttat catcagtgct ctca 24
<210> 197
<211> 26
<212> DNA
<213> Homo sapiens
<400> 197
ctttgtccag atgaactctt tgagaa 26
<210> 198
<211> 22
<212> DNA
<213> Homo sapiens
<400> 198
aacttcgctc acaggatctt ca 22
<210> 199
<211> 22
<212> DNA
<213> Homo sapiens
<400> 199
ggtggaggtg ttttacttct gc 22
<210> 200
<211> 28
<212> DNA
<213> Homo sapiens
<400> 200
tcatcatctg aatcatctaa taggtcct 28
<210> 201
<211> 26
<212> DNA
<213> Homo sapiens
<400> 201
ctgtaggaat ggtatctcgt ttttca 26
<210> 202
<211> 26
<212> DNA
<213> Homo sapiens
<400> 202
tctttgttgt ctgagaaaac tctctc 26
<210> 203
<211> 22
<212> DNA
<213> Homo sapiens
<400> 203
gctgacctcg atttattggc tg 22
<210> 204
<211> 24
<212> DNA
<213> Homo sapiens
<400> 204
gcaacaggtc atcttcagag tcaa 24
<210> 205
<211> 26
<212> DNA
<213> Homo sapiens
<400> 205
aattcgtggg cctttattac ttgtaa 26
<210> 206
<211> 22
<212> DNA
<213> Homo sapiens
<400> 206
gatttcacag atggcttggc tc 22
<210> 207
<211> 26
<212> DNA
<213> Homo sapiens
<400> 207
tggctccatt accattattc atctga 26
<210> 208
<211> 22
<212> DNA
<213> Homo sapiens
<400> 208
gctttgctgg tcttccatca tt 22
<210> 209
<211> 22
<212> DNA
<213> Homo sapiens
<400> 209
attcagcacc atttgtagca cc 22
<210> 210
<211> 22
<212> DNA
<213> Homo sapiens
<400> 210
ctgaatgggg tacgttccac ta 22
<210> 211
<211> 30
<212> DNA
<213> Homo sapiens
<400> 211
tcatttgaaa caaaatgtct atatagcagt 30
<210> 212
<211> 26
<212> DNA
<213> Homo sapiens
<400> 212
cgtgtttgtc aagtcatgga tttttc 26
<210> 213
<211> 35
<212> DNA
<213> Homo sapiens
<400> 213
acataatcag aaaaataatc ttgtaaaatg tctga 35
<210> 214
<211> 25
<212> DNA
<213> Homo sapiens
<400> 214
ctgacgactc tttttcagat gtgga 25
<210> 215
<211> 23
<212> DNA
<213> Homo sapiens
<400> 215
aaagacgtgt ccatttccag act 23
<210> 216
<211> 23
<212> DNA
<213> Homo sapiens
<400> 216
agagtgagac gctgtctgaa aat 23
<210> 217
<211> 23
<212> DNA
<213> Homo sapiens
<400> 217
tgtcacctgg tgttttgttt tca 23
<210> 218
<211> 32
<212> DNA
<213> Homo sapiens
<400> 218
cttgagctgt gtaattccta taatattaaa gt 32
<210> 219
<211> 24
<212> DNA
<213> Homo sapiens
<400> 219
gtcttgggaa atgcaaaaac agtt 24
<210> 220
<211> 27
<212> DNA
<213> Homo sapiens
<400> 220
attatttaca gtgttgagtc atttccc 27
<210> 221
<211> 18
<212> DNA
<213> Homo sapiens
<400> 221
gttgtcccca catgcccc 18
<210> 222
<211> 18
<212> DNA
<213> Homo sapiens
<400> 222
ctgtgcgagc ctcccctc 18
<210> 223
<211> 18
<212> DNA
<213> Homo sapiens
<400> 223
caagacgcag aggcccct 18
<210> 224
<211> 18
<212> DNA
<213> Homo sapiens
<400> 224
ctgatgcccc tcgctctg 18
<210> 225
<211> 18
<212> DNA
<213> Homo sapiens
<400> 225
caccacgccc tgctctag 18
<210> 226
<211> 23
<212> DNA
<213> Homo sapiens
<400> 226
acaacaggat tctcagggaa tgg 23
<210> 227
<211> 25
<212> DNA
<213> Homo sapiens
<400> 227
gaacatgtag gaaacacaag gaagt 25
<210> 228
<211> 25
<212> DNA
<213> Homo sapiens
<400> 228
cattctctct tgttactcat gccac 25
<210> 229
<211> 22
<212> DNA
<213> Homo sapiens
<400> 229
gagtgtcata gggcagagtc ac 22
<210> 230
<211> 22
<212> DNA
<213> Homo sapiens
<400> 230
cacagagggg ccaaagagtt ag 22
<210> 231
<211> 22
<212> DNA
<213> Homo sapiens
<400> 231
tgtgggtagg atcagaggtc aa 22
<210> 232
<211> 21
<212> DNA
<213> Homo sapiens
<400> 232
catgaggaca ctgctgacct g 21
<210> 233
<211> 22
<212> DNA
<213> Homo sapiens
<400> 233
acgtatcaca atcccttccc ag 22
<210> 234
<211> 26
<212> DNA
<213> Homo sapiens
<400> 234
agcaagacca attattaaag gtgaca 26
<210> 235
<211> 22
<212> DNA
<213> Homo sapiens
<400> 235
aaggccatct gtacctgttc ac 22
<210> 236
<211> 23
<212> DNA
<213> Homo sapiens
<400> 236
caggctacct agaattgaac acg 23
<210> 237
<211> 26
<212> DNA
<213> Homo sapiens
<400> 237
gtttctttca tcaagagctc aaacct 26
<210> 238
<211> 26
<212> DNA
<213> Homo sapiens
<400> 238
cagagccctg cttaaattgg cttatt 26
<210> 239
<211> 23
<212> DNA
<213> Homo sapiens
<400> 239
tcttgtccag caaccatttt tgt 23
<210> 240
<211> 20
<212> DNA
<213> Homo sapiens
<400> 240
ctctggctgc tgaggagaag 20
<210> 241
<211> 27
<212> DNA
<213> Homo sapiens
<400> 241
ttagtttgat tgctgcatat ttcagat 27
<210> 242
<211> 26
<212> DNA
<213> Homo sapiens
<400> 242
tgtgtcacat tataaagatt caggca 26
<210> 243
<211> 26
<212> DNA
<213> Homo sapiens
<400> 243
ccaatggcta agtgaagatg acaatc 26
<210> 244
<211> 26
<212> DNA
<213> Homo sapiens
<400> 244
ggcatttcct gtgaaataat actggt 26
<210> 245
<211> 26
<212> DNA
<213> Homo sapiens
<400> 245
tggaagtcta tgtgatcaag aaatcg 26
<210> 246
<211> 26
<212> DNA
<213> Homo sapiens
<400> 246
caaaattgct tcagaaattg gagaca 26
<210> 247
<211> 26
<212> DNA
<213> Homo sapiens
<400> 247
tgtcttgcat aatgtgacac atgaat 26
<210> 248
<211> 33
<212> DNA
<213> Homo sapiens
<400> 248
tttctgggaa tagaagctta taaaaattta aaa 33
<210> 249
<211> 33
<212> DNA
<213> Homo sapiens
<400> 249
acatctatga atgtaccatg ttaatgtctt ctt 33
<210> 250
<211> 27
<212> DNA
<213> Homo sapiens
<400> 250
agttcttaga cattgcataa gcttgtt 27
<210> 251
<211> 26
<212> DNA
<213> Homo sapiens
<400> 251
gagtgcaagt gaaagcctta tatctt 26
<210> 252
<211> 29
<212> DNA
<213> Homo sapiens
<400> 252
ttaagcatgc tatacaatct gaactaaaa 29
<210> 253
<211> 32
<212> DNA
<213> Homo sapiens
<400> 253
tggtttttaa tgtatggaat ttttctttat ga 32
<210> 254
<211> 22
<212> DNA
<213> Homo sapiens
<400> 254
tgtagggagg atgggaagag at 22
<210> 255
<211> 22
<212> DNA
<213> Homo sapiens
<400> 255
gaagtcggaa cacaaggaag ga 22
<210> 256
<211> 22
<212> DNA
<213> Homo sapiens
<400> 256
cttgactgac cacgcctttc tt 22
<210> 257
<211> 27
<212> DNA
<213> Homo sapiens
<400> 257
tgaaatcagt tatttttcag tttggca 27
<210> 258
<211> 26
<212> DNA
<213> Homo sapiens
<400> 258
tggaagactg agttatagtc aactga 26
<210> 259
<211> 29
<212> DNA
<213> Homo sapiens
<400> 259
aatcaaacac tgaatattct gattctcaa 29
<210> 260
<211> 26
<212> DNA
<213> Homo sapiens
<400> 260
gtaattatat gtggccatgg ttggtt 26
<210> 261
<211> 35
<212> DNA
<213> Homo sapiens
<400> 261
ttaaaagcat atatgtctgt ttagataatc ttttt 35
<210> 262
<211> 22
<212> DNA
<213> Homo sapiens
<400> 262
cgcattttct tcaaccagga gg 22
<210> 263
<211> 26
<212> DNA
<213> Homo sapiens
<400> 263
cagctaatac agtgcttgaa catgta 26
<210> 264
<211> 22
<212> DNA
<213> Homo sapiens
<400> 264
ttgtgggtgt taaaatgtcc gc 22
<210> 265
<211> 33
<212> DNA
<213> Homo sapiens
<400> 265
taaacattta atgtaggtga atctgttatc act 33
<210> 266
<211> 26
<212> DNA
<213> Homo sapiens
<400> 266
tagttttctt ggtttggatt gggaag 26
<210> 267
<211> 26
<212> DNA
<213> Homo sapiens
<400> 267
ccagcagatt caagcttttt aaatgg 26
<210> 268
<211> 28
<212> DNA
<213> Homo sapiens
<400> 268
tgaaaacagt aaaatttaag tgggagga 28
<210> 269
<211> 24
<212> DNA
<213> Homo sapiens
<400> 269
tgtttcacgt agtacacatt gctt 24
<210> 270
<211> 24
<212> DNA
<213> Homo sapiens
<400> 270
gagcacctgt tccatatgta cgac 24
<210> 271
<211> 22
<212> DNA
<213> Homo sapiens
<400> 271
tgtggcatat ccttcccaat gt 22
<210> 272
<211> 26
<212> DNA
<213> Homo sapiens
<400> 272
taaattgctg tctcttctca tgctgt 26
<210> 273
<211> 26
<212> DNA
<213> Homo sapiens
<400> 273
tgcttcaata ttaatgccag aagact 26
<210> 274
<211> 33
<212> DNA
<213> Homo sapiens
<400> 274
ggttttccaa attttgattt gtttttaaat act 33
<210> 275
<211> 22
<212> DNA
<213> Homo sapiens
<400> 275
aaagctctag gaaggaaacg cc 22
<210> 276
<211> 22
<212> DNA
<213> Homo sapiens
<400> 276
atgagctgta ccacatggat gc 22
<210> 277
<211> 22
<212> DNA
<213> Homo sapiens
<400> 277
gccatactcg tgcatatggt gt 22
<210> 278
<211> 25
<212> DNA
<213> Homo sapiens
<400> 278
gagagaaaag tgaattggcc ctctc 25
<210> 279
<211> 25
<212> DNA
<213> Homo sapiens
<400> 279
ctagatgcca tagaagacct catgg 25
<210> 280
<211> 22
<212> DNA
<213> Homo sapiens
<400> 280
tgaccatgaa aaggctcgaa ag 22
<210> 281
<211> 26
<212> DNA
<213> Homo sapiens
<400> 281
aatctcataa atgctgaaga acggag 26
<210> 282
<211> 22
<212> DNA
<213> Homo sapiens
<400> 282
gatcacgcca tccttgcatt ac 22
<210> 283
<211> 33
<212> DNA
<213> Homo sapiens
<400> 283
cctagaagat gaatttatgt aatatgattt gca 33
<210> 284
<211> 24
<212> DNA
<213> Homo sapiens
<400> 284
ttgctagcac atgtatcgct aata 24
<210> 285
<211> 22
<212> DNA
<213> Homo sapiens
<400> 285
acttccgttg agcatctaga cg 22
<210> 286
<211> 27
<212> DNA
<213> Homo sapiens
<400> 286
tcctggatta aatcaagaaa atgggaa 27
<210> 287
<211> 28
<212> DNA
<213> Homo sapiens
<400> 287
tgttgatatg attttctctt ttcccctt 28
<210> 288
<211> 22
<212> DNA
<213> Homo sapiens
<400> 288
actaaaaggg ggctctgaca tc 22
<210> 289
<211> 24
<212> DNA
<213> Homo sapiens
<400> 289
agtttatggg aaggaacctt gtgt 24
<210> 290
<211> 22
<212> DNA
<213> Homo sapiens
<400> 290
tctccccctc ccactatcta ag 22
<210> 291
<211> 22
<212> DNA
<213> Homo sapiens
<400> 291
gggaacagaa gcttgatgca tt 22
<210> 292
<211> 32
<212> DNA
<213> Homo sapiens
<400> 292
agtttaaaaa caagaataat aatgatctgc ac 32
<210> 293
<211> 22
<212> DNA
<213> Homo sapiens
<400> 293
cccattttgt cccaactggt tg 22
<210> 294
<211> 22
<212> DNA
<213> Homo sapiens
<400> 294
gagtggcaga taggagcaca ag 22
<210> 295
<211> 23
<212> DNA
<213> Homo sapiens
<400> 295
caaacaggga ggcttatgac atc 23
<210> 296
<211> 30
<212> DNA
<213> Homo sapiens
<400> 296
agtagtgaat ttcaaaatcc tttttaacct 30
<210> 297
<211> 29
<212> DNA
<213> Homo sapiens
<400> 297
gcttaaagca attgttgtat aaaaacttg 29
<210> 298
<211> 26
<212> DNA
<213> Homo sapiens
<400> 298
tgcttttact gattaacgta aataca 26
<210> 299
<211> 25
<212> DNA
<213> Homo sapiens
<400> 299
ctaagaaagc ctacaccatt tttgc 25
<210> 300
<211> 26
<212> DNA
<213> Homo sapiens
<400> 300
ttcatcactt aattggtttt tggctt 26
<210> 301
<211> 26
<212> DNA
<213> Homo sapiens
<400> 301
cgtgaaatcc gagtccttca tctttt 26
<210> 302
<211> 26
<212> DNA
<213> Homo sapiens
<400> 302
ccaacttggt accagtttgt tttatt 26
<210> 303
<211> 27
<212> DNA
<213> Homo sapiens
<400> 303
agatttctat tcttactgct agcatta 27
<210> 304
<211> 26
<212> DNA
<213> Homo sapiens
<400> 304
cacattgcac tgagaataaa gctgat 26
<210> 305
<211> 19
<212> DNA
<213> Homo sapiens
<400> 305
gggacatggg ggcagttag 19
<210> 306
<211> 22
<212> DNA
<213> Homo sapiens
<400> 306
gcaacatgac tgtcctttca cc 22
<210> 307
<211> 26
<212> DNA
<213> Homo sapiens
<400> 307
acacttacaa tttcactaag tcggaa 26
<210> 308
<211> 26
<212> DNA
<213> Homo sapiens
<400> 308
gcaagaccca aacacataat agaaga 26
<210> 309
<211> 22
<212> DNA
<213> Homo sapiens
<400> 309
aacgtcatgt ggatcagcct at 22
<210> 310
<211> 22
<212> DNA
<213> Homo sapiens
<400> 310
gacgacacag gaagcagatt ct 22
<210> 311
<211> 26
<212> DNA
<213> Homo sapiens
<400> 311
gtggaatggt aagtggcatt ataagc 26
<210> 312
<211> 26
<212> DNA
<213> Homo sapiens
<400> 312
acaatgggaa tgaaacagaa tcagag 26
<210> 313
<211> 23
<212> DNA
<213> Homo sapiens
<400> 313
tctaacaatc gaatcccctc caa 23
<210> 314
<211> 27
<212> DNA
<213> Homo sapiens
<400> 314
accacaaaat actgaatata ggacacg 27
<210> 315
<211> 22
<212> DNA
<213> Homo sapiens
<400> 315
taccagccac acagaactaa cc 22
<210> 316
<211> 25
<212> DNA
<213> Homo sapiens
<400> 316
agatacccca gtttgtttct caaga 25
<210> 317
<211> 22
<212> DNA
<213> Homo sapiens
<400> 317
caggaatctc tctgggatca cc 22
<210> 318
<211> 24
<212> DNA
<213> Homo sapiens
<400> 318
ccaaaagccc tagtgaaggt caaa 24
<210> 319
<211> 25
<212> DNA
<213> Homo sapiens
<400> 319
accaaacaaa caggtttatc caaga 25
<210> 320
<211> 24
<212> DNA
<213> Homo sapiens
<400> 320
gtctctatcc acacattcgt ctgt 24
<210> 321
<211> 31
<212> DNA
<213> Homo sapiens
<400> 321
ggaacatgga gaaaaataaa agaaaatgaa t 31
<210> 322
<211> 24
<212> DNA
<213> Homo sapiens
<400> 322
aggacaaaat aatcctgtcc ctgt 24
<210> 323
<211> 27
<212> DNA
<213> Homo sapiens
<400> 323
tgtctttcaa aacataaaaa tctgcga 27
<210> 324
<211> 26
<212> DNA
<213> Homo sapiens
<400> 324
cagagtgctg ggattacaga cgtgag 26
<210> 325
<211> 19
<212> DNA
<213> Homo sapiens
<400> 325
tgcgcaacag agcaagact 19
<210> 326
<211> 26
<212> DNA
<213> Homo sapiens
<400> 326
cgaaatttac atccggtatc ttcctg 26
<210> 327
<211> 26
<212> DNA
<213> Homo sapiens
<400> 327
ctagaagaca gcatacccct tttctg 26
<210> 328
<211> 23
<212> DNA
<213> Homo sapiens
<400> 328
cagagcaagt ccctgtctca aaa 23
<210> 329
<211> 22
<212> DNA
<213> Homo sapiens
<400> 329
attccagtca tagcagagct gt 22
<210> 330
<211> 27
<212> DNA
<213> Homo sapiens
<400> 330
acacgaaact attagcctta gaatcac 27
<210> 331
<211> 26
<212> DNA
<213> Homo sapiens
<400> 331
aaatcttttg ctcatgtgca ttaacc 26
<210> 332
<211> 23
<212> DNA
<213> Homo sapiens
<400> 332
tgcatttccc aagacagtgt tac 23
<210> 333
<211> 26
<212> DNA
<213> Homo sapiens
<400> 333
ctcttcgtct tttctaaccg tgcagc 26
<210> 334
<211> 18
<212> DNA
<213> Homo sapiens
<400> 334
tcctcgtgtc ccactccc 18
<210> 335
<211> 20
<212> DNA
<213> Homo sapiens
<400> 335
tgcttttagg agctccgtcc 20
<210> 336
<211> 18
<212> DNA
<213> Homo sapiens
<400> 336
ccttttctgg cccccgtg 18
<210> 337
<211> 18
<212> DNA
<213> Homo sapiens
<400> 337
gcagctgcaa aggggacc 18
<210> 338
<211> 22
<212> DNA
<213> Homo sapiens
<400> 338
gaatgccgtg ggtctcaaag ag 22
<210> 339
<211> 18
<212> DNA
<213> Homo sapiens
<400> 339
ctggtgccag cctgacag 18
<210> 340
<211> 22
<212> DNA
<213> Homo sapiens
<400> 340
agtggagaat gttcacccag ac 22
<210> 341
<211> 22
<212> DNA
<213> Homo sapiens
<400> 341
tagcctcaaa agccaacatc ct 22
<210> 342
<211> 22
<212> DNA
<213> Homo sapiens
<400> 342
ccgggttctg cagaatctta ct 22
<210> 343
<211> 22
<212> DNA
<213> Homo sapiens
<400> 343
aaagaaggga acactgctgt ga 22
<210> 344
<211> 22
<212> DNA
<213> Homo sapiens
<400> 344
ctaccacctg attggagtgc aa 22
<210> 345
<211> 22
<212> DNA
<213> Homo sapiens
<400> 345
gtcaagggtg aaggtggtag ag 22
<210> 346
<211> 18
<212> DNA
<213> Homo sapiens
<400> 346
tgcctcccac ccactgtc 18
<210> 347
<211> 19
<212> DNA
<213> Homo sapiens
<400> 347
acggaagttc gacccatcg 19
<210> 348
<211> 27
<212> DNA
<213> Homo sapiens
<400> 348
gaaacaatga gcttttcaga aatgatt 27
<210> 349
<211> 29
<212> DNA
<213> Homo sapiens
<400> 349
aagacatatc agtttaaaat accaaacca 29
<210> 350
<211> 27
<212> DNA
<213> Homo sapiens
<400> 350
aggataatgg tatagagaac ctcaagg 27
<210> 351
<211> 26
<212> DNA
<213> Homo sapiens
<400> 351
ttctccctag cctatctctg atgata 26
<210> 352
<211> 23
<212> DNA
<213> Homo sapiens
<400> 352
ttatttttgg ccctcaactt gga 23
<210> 353
<211> 26
<212> DNA
<213> Homo sapiens
<400> 353
aagtgattcg taaatgccac caaata 26
<210> 354
<211> 22
<212> DNA
<213> Homo sapiens
<400> 354
tttccatcct gcagaagaag cc 22
<210> 355
<211> 28
<212> DNA
<213> Homo sapiens
<400> 355
aagaaaatct gtcttttggt ttttcttg 28
<210> 356
<211> 27
<212> DNA
<213> Homo sapiens
<400> 356
acctgttaag tttgtatgca acatttc 27
<210> 357
<211> 26
<212> DNA
<213> Homo sapiens
<400> 357
acccagttac catagcaatt tagtga 26
<210> 358
<211> 26
<212> DNA
<213> Homo sapiens
<400> 358
taacataggt gacagatttt cttttt 26
<210> 359
<211> 26
<212> DNA
<213> Homo sapiens
<400> 359
agatgagtca tatttgtggg ttttca 26
<210> 360
<211> 22
<212> DNA
<213> Homo sapiens
<400> 360
atgtgccata gacaaggtgg ag 22
<210> 361
<211> 31
<212> DNA
<213> Homo sapiens
<400> 361
atctttggtt taaatttaca ttctctgttt t 31
<210> 362
<211> 22
<212> DNA
<213> Homo sapiens
<400> 362
atgtgcatga ctttgaggga ca 22
<210> 363
<211> 25
<212> DNA
<213> Homo sapiens
<400> 363
agtaggcagc ctttataaaa gcaaa 25
<210> 364
<211> 26
<212> DNA
<213> Homo sapiens
<400> 364
ggtaggagag acatttaagg ttcctt 26
<210> 365
<211> 26
<212> DNA
<213> Homo sapiens
<400> 365
gttggaatgt aaaggtgaag gtgatg 26
<210> 366
<211> 22
<212> DNA
<213> Homo sapiens
<400> 366
tgttgatgac cttcgtcgct ta 22
<210> 367
<211> 18
<212> DNA
<213> Homo sapiens
<400> 367
aagcgggcca agctcatc 18
<210> 368
<211> 20
<212> DNA
<213> Homo sapiens
<400> 368
atccaggcca tcatcctgac 20
<210> 369
<211> 23
<212> DNA
<213> Homo sapiens
<400> 369
aacttagctg ggacatgaga gtt 23
<210> 370
<211> 22
<212> DNA
<213> Homo sapiens
<400> 370
ttgggagaag agcaaaacct ga 22
<210> 371
<211> 22
<212> DNA
<213> Homo sapiens
<400> 371
acagttttag accctgagct gt 22
<210> 372
<211> 26
<212> DNA
<213> Homo sapiens
<400> 372
aagaaaatgg acctgacagt aaatgg 26
<210> 373
<211> 35
<212> DNA
<213> Homo sapiens
<400> 373
actattttca gaatgaacaa aagattgaaa aatta 35
<210> 374
<211> 19
<212> DNA
<213> Homo sapiens
<400> 374
ggcttcgtgc gcttctttc 19
<210> 375
<211> 31
<212> DNA
<213> Homo sapiens
<400> 375
agtttggatt tttccttttt gcttataaaa t 31
<210> 376
<211> 28
<212> DNA
<213> Homo sapiens
<400> 376
cacaaaagac atttatcagg acctcaac 28
<210> 377
<211> 26
<212> DNA
<213> Homo sapiens
<400> 377
attttaggtt gcagtttcat cactgt 26
<210> 378
<211> 23
<212> DNA
<213> Homo sapiens
<400> 378
aggttttcac taatgagctt gcc 23
<210> 379
<211> 22
<212> DNA
<213> Homo sapiens
<400> 379
acttcgtcga ttcccagatc tt 22
<210> 380
<211> 26
<212> DNA
<213> Homo sapiens
<400> 380
ccttttggat caaatgatgc ttgttt 26
<210> 381
<211> 32
<212> DNA
<213> Homo sapiens
<400> 381
tggtagtagg tatttatgga atactttttc tt 32
<210> 382
<211> 27
<212> DNA
<213> Homo sapiens
<400> 382
tcagtattcc tgtgtacatt ttctgtt 27
<210> 383
<211> 29
<212> DNA
<213> Homo sapiens
<400> 383
agaaagaagt ttaaaatctt gctttctga 29
<210> 384
<211> 22
<212> DNA
<213> Homo sapiens
<400> 384
cgatggattt gggttagcat gg 22
<210> 385
<211> 35
<212> DNA
<213> Homo sapiens
<400> 385
cttttagaaa agatatttta attactaatg ggaca 35
<210> 386
<211> 22
<212> DNA
<213> Homo sapiens
<400> 386
gtggccttgt ctggtttaca ac 22
<210> 387
<211> 21
<212> DNA
<213> Homo sapiens
<400> 387
gcacccggcc cttattgttt a 21
<210> 388
<211> 22
<212> DNA
<213> Homo sapiens
<400> 388
gagtgacatt ggtggctctg at 22
<210> 389
<211> 22
<212> DNA
<213> Homo sapiens
<400> 389
gagaagagat gagcacagga gg 22
<210> 390
<211> 22
<212> DNA
<213> Homo sapiens
<400> 390
gcacgatgta gaaagatggc ac 22
<210> 391
<211> 23
<212> DNA
<213> Homo sapiens
<400> 391
ctgttctctt caggaaggtc tga 23
<210> 392
<211> 24
<212> DNA
<213> Homo sapiens
<400> 392
tgcagtgaca ttaaacaact tgga 24
<210> 393
<211> 24
<212> DNA
<213> Homo sapiens
<400> 393
tgattttctt tctgctctgg aagg 24
<210> 394
<211> 22
<212> DNA
<213> Homo sapiens
<400> 394
accagctgga aattcctgag aa 22
<210> 395
<211> 22
<212> DNA
<213> Homo sapiens
<400> 395
taaaaccccc aaacgatgaa gc 22
<210> 396
<211> 22
<212> DNA
<213> Homo sapiens
<400> 396
gcctagctct tacgtaaggg tt 22
<210> 397
<211> 30
<212> DNA
<213> Homo sapiens
<400> 397
gttttaattc ctttgagtta cttccttatg 30
<210> 398
<211> 23
<212> DNA
<213> Homo sapiens
<400> 398
acagtaaaag gggaagggat gat 23
<210> 399
<211> 24
<212> DNA
<213> Homo sapiens
<400> 399
tggagtttgt tatcattgct tggc 24
<210> 400
<211> 22
<212> DNA
<213> Homo sapiens
<400> 400
gtgacccagc agtgagtttt tc 22
<210> 401
<211> 22
<212> DNA
<213> Homo sapiens
<400> 401
cttaagacct cgcttttgcc ag 22
<210> 402
<211> 22
<212> DNA
<213> Homo sapiens
<400> 402
agtttgctgg tggagataag gt 22
<210> 403
<211> 22
<212> DNA
<213> Homo sapiens
<400> 403
acccctcagg acagttttga ac 22
<210> 404
<211> 26
<212> DNA
<213> Homo sapiens
<400> 404
acttcttatt ctgagtctct ccacta 26
<210> 405
<211> 22
<212> DNA
<213> Homo sapiens
<400> 405
ggctgccaaa aatcagagct tg 22
<210> 406
<211> 22
<212> DNA
<213> Homo sapiens
<400> 406
tgaagtgggg ttggtaggat tc 22
<210> 407
<211> 24
<212> DNA
<213> Homo sapiens
<400> 407
cgttaaagct tgctccttca tgtt 24
<210> 408
<211> 25
<212> DNA
<213> Homo sapiens
<400> 408
agctttaagt agtctgtgat ctccg 25
<210> 409
<211> 21
<212> DNA
<213> Homo sapiens
<400> 409
ttggtgagga aggtgaagca c 21
<210> 410
<211> 30
<212> DNA
<213> Homo sapiens
<400> 410
tcagcatact taaatgtcaa gaaatacaga 30
<210> 411
<211> 26
<212> DNA
<213> Homo sapiens
<400> 411
gctcttctgc agtctttatt agcatt 26
<210> 412
<211> 26
<212> DNA
<213> Homo sapiens
<400> 412
gaatgatttg acataaccct gagctt 26
<210> 413
<211> 28
<212> DNA
<213> Homo sapiens
<400> 413
cacttcattt ggagtacctt aacatgat 28
<210> 414
<211> 22
<212> DNA
<213> Homo sapiens
<400> 414
cctcatccag cttttacatg gc 22
<210> 415
<211> 25
<212> DNA
<213> Homo sapiens
<400> 415
agggttatat tagtgatccc tgcat 25
<210> 416
<211> 23
<212> DNA
<213> Homo sapiens
<400> 416
aagcttggct tcaagttgtc ttt 23
<210> 417
<211> 28
<212> DNA
<213> Homo sapiens
<400> 417
acaaattcca actctaatta gatgaccc 28
<210> 418
<211> 24
<212> DNA
<213> Homo sapiens
<400> 418
tgttactgca tacacattgt gacc 24
<210> 419
<211> 23
<212> DNA
<213> Homo sapiens
<400> 419
ggaaacaaaa agccctagaa gca 23
<210> 420
<211> 23
<212> DNA
<213> Homo sapiens
<400> 420
aggaacttct tcaaagcgag gtt 23
<210> 421
<211> 22
<212> DNA
<213> Homo sapiens
<400> 421
cagccgacct agcccataaa at 22
<210> 422
<211> 26
<212> DNA
<213> Homo sapiens
<400> 422
tcgagtgggt tctaatcatg gaatta 26
<210> 423
<211> 22
<212> DNA
<213> Homo sapiens
<400> 423
agagtagaag tggtcagcct ca 22
<210> 424
<211> 22
<212> DNA
<213> Homo sapiens
<400> 424
agcgaaatct ccctccaaaa gt 22
<210> 425
<211> 24
<212> DNA
<213> Homo sapiens
<400> 425
tccaggttct tccagatgct gata 24
<210> 426
<211> 24
<212> DNA
<213> Homo sapiens
<400> 426
cccagactgc ttcaaaatta cctc 24
<210> 427
<211> 24
<212> DNA
<213> Homo sapiens
<400> 427
aagcagagga aggtgatatt cttg 24
<210> 428
<211> 23
<212> DNA
<213> Homo sapiens
<400> 428
gattcacctc atcattacac gcc 23
<210> 429
<211> 29
<212> DNA
<213> Homo sapiens
<400> 429
actgatgaaa agttacagaa ttttgctat 29
<210> 430
<211> 27
<212> DNA
<213> Homo sapiens
<400> 430
tgatttgaaa gatatacaga gaccaga 27
<210> 431
<211> 27
<212> DNA
<213> Homo sapiens
<400> 431
agtttgatta ctggaaaagt tcgatct 27
<210> 432
<211> 24
<212> DNA
<213> Homo sapiens
<400> 432
tagacctata cagtctcctg gccg 24
<210> 433
<211> 22
<212> DNA
<213> Homo sapiens
<400> 433
acgccagtca actttcatca aa 22
<210> 434
<211> 22
<212> DNA
<213> Homo sapiens
<400> 434
cctcgagtaa gcacttggag aa 22
<210> 435
<211> 22
<212> DNA
<213> Homo sapiens
<400> 435
ggtaatactc ccccggtgat tg 22
<210> 436
<211> 26
<212> DNA
<213> Homo sapiens
<400> 436
tccttttaat tacaacccaa gcccta 26
<210> 437
<211> 21
<212> DNA
<213> Homo sapiens
<400> 437
gagaccttcc tcgactgcaa g 21
<210> 438
<211> 26
<212> DNA
<213> Homo sapiens
<400> 438
ttagatcttc aatttgaggg ggagtc 26
<210> 439
<211> 23
<212> DNA
<213> Homo sapiens
<400> 439
atagaaaagt ccaggggcac aac 23
<210> 440
<211> 23
<212> DNA
<213> Homo sapiens
<400> 440
gaacccctca gaatccacgg aag 23
<210> 441
<211> 26
<212> DNA
<213> Homo sapiens
<400> 441
ctgactgaca tttagcttgt tgacat 26
<210> 442
<211> 27
<212> DNA
<213> Homo sapiens
<400> 442
caaaggcata aagaacaaac taacaca 27
<210> 443
<211> 25
<212> DNA
<213> Homo sapiens
<400> 443
aatcactaga gcaataagag gcgtt 25
<210> 444
<211> 22
<212> DNA
<213> Homo sapiens
<400> 444
tctaaggggt caagtgagtg ga 22
<210> 445
<211> 26
<212> DNA
<213> Homo sapiens
<400> 445
attcacagat catttcttgt ggctta 26
<210> 446
<211> 18
<212> DNA
<213> Homo sapiens
<400> 446
gttgcgcccc tcagctca 18
<210> 447
<211> 20
<212> DNA
<213> Homo sapiens
<400> 447
gctgagtgat gccaacaagg 20
<210> 448
<211> 19
<212> DNA
<213> Homo sapiens
<400> 448
ccgaggtagg cacgtgcta 19
<210> 449
<211> 20
<212> DNA
<213> Homo sapiens
<400> 449
ttctactgcc tccaacacgg 20
<210> 450
<211> 19
<212> DNA
<213> Homo sapiens
<400> 450
cctgacaaca gaggctggg 19

Claims (10)

1. a kind of multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection, it is characterised in that more than first Weight PCR primer combination and/or the combination of the second multiple PCR primer；Wherein, the first multiple PCR primer combination contains 112 pairs PCR primer pair, and the sequence of 112 pairs of PCR primers pair is respectively such as SEQ ID NO:1~SEQ ID NO:Shown in 224；Described The combination of one multiple PCR primer contains 113 pairs of PCR primers pair, and the sequence of 113 pairs of PCR primers pair is respectively such as SEQ ID NO: 225~SEQ ID NO:Shown in 450.
2. the multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection as claimed in claim 1, its feature exist In each pair of primer equimolar amounts mixing in the first multiple PCR primer combination.
3. the multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection as claimed in claim 1, its feature exist In each pair of primer equimolar amounts mixing in the second multiple PCR primer combination.
4. such as the multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection according to any one of claims 1 to 3 The application united in the detection reagent for preparing the detection straight colon cancer susceptibility gene of heredity.
5. a kind of heredity susceptibility gene of colorectal cancer detection kit, it is characterised in that containing in such as claims 1 to 3 The multiple PCR primer system of heredity susceptibility gene of colorectal cancer detection described in any one.
6. heredity susceptibility gene of colorectal cancer detection kit as claimed in claim 5, it is characterised in that also include DNA extracts reagents, PCR amplifing reagents, and/or DNA purified reagents.
7. a kind of detection method of heredity susceptibility gene of colorectal cancer, it is characterised in that comprise the following steps：
Use the multiple PCR primer of heredity susceptibility gene of colorectal cancer detection such as according to any one of claims 1 to 3 System, multiplexed PCR amplification is carried out by template of the genomic DNA of acquisition；
Sequencing library is built to the product of multiplexed PCR amplification；
High-flux sequence is carried out to the sequencing library of structure, and sequencing result is compared onto human genome reference sequences.
8. the detection method of heredity susceptibility gene of colorectal cancer as claimed in claim 7, it is characterised in that the structure is surveyed Preface storehouse includes successively：End reparation is carried out to the product of multiplexed PCR amplification, end adds A, connection sequence measuring joints, amplified library, Library after amplification is purified and fragment is screened.
9. the detection method of heredity susceptibility gene of colorectal cancer as claimed in claim 7, it is characterised in that mankind's base Because group reference sequences are hg19.
10. the detection method of heredity susceptibility gene of colorectal cancer as claimed in claim 7, it is characterised in that also include pair Comparison result carries out the step of Mutation Screening analysis and mutation progress functional annotation to screening.
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CN112831563A (en) * 2021-02-08 2021-05-25 复旦大学附属肿瘤医院 Detection panel and detection kit for hereditary colorectal cancer
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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110684848A (en) * 2019-10-25 2020-01-14 广州万德基因医学科技有限公司 Multiple PCR primer group and kit for genetic tumor germ line mutation detection
CN112831563A (en) * 2021-02-08 2021-05-25 复旦大学附属肿瘤医院 Detection panel and detection kit for hereditary colorectal cancer
CN113637755A (en) * 2021-08-17 2021-11-12 珠海市人民医院 Primer composition for detecting cancer susceptibility gene and application thereof
CN117844933A (en) * 2024-03-07 2024-04-09 上海复迪生生命科学有限公司 Multiplex PCR primer group for detecting lung tumor related gene variation and application thereof
CN117844933B (en) * 2024-03-07 2024-06-11 上海复迪生生命科学有限公司 Multiplex PCR primer group for detecting lung tumor related gene variation and application thereof

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