WO2017120750A1 - Puce à adn d'une région non codante des snp à l'intérieur de la plage du génome entier de la population d'asie de l'est - Google Patents

Puce à adn d'une région non codante des snp à l'intérieur de la plage du génome entier de la population d'asie de l'est Download PDF

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Publication number
WO2017120750A1
WO2017120750A1 PCT/CN2016/070641 CN2016070641W WO2017120750A1 WO 2017120750 A1 WO2017120750 A1 WO 2017120750A1 CN 2016070641 W CN2016070641 W CN 2016070641W WO 2017120750 A1 WO2017120750 A1 WO 2017120750A1
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Prior art keywords
snps
coding
dna chip
east asian
whole genome
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PCT/CN2016/070641
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English (en)
Chinese (zh)
Inventor
陈小伟
陈润生
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中国科学院生物物理研究所
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Priority to PCT/CN2016/070641 priority Critical patent/WO2017120750A1/fr
Priority to CN201680000526.7A priority patent/CN107018668B/zh
Publication of WO2017120750A1 publication Critical patent/WO2017120750A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/156—Polymorphic or mutational markers

Definitions

  • the present invention relates to the fields of molecular biology, functional genomics, bioinformatics, and molecular diagnostics, and more particularly to a DNA chip for SNPs of non-coding regions within the genome-wide range of East Asian populations.
  • the NONCODE database is the first database to collect non-coding RNA sequences. It has been updated to the fourth edition and is the most authoritative database in the non-coding field. Beginning with the third edition, the NONCODE database began collecting long-chain non-coding RNA sequences.
  • SNPs Single nucleotide polymorphisms
  • SNP Single nucleotide polymorphisms
  • MAF minimum equipotential frequency
  • SNPs can be divided into gene coding region SNP, gene non-coding region SNP and intergenic region SNP.
  • Protein coding region SNP loci are further divided into two types: synonymous and non-synonymous. Due to the annexation of codons, synonymous SNPs do not alter protein sequences, while non-synonymous SNPs can alter protein sequences. Non-synonymous SNPs are further divided into missense and nonsense. SNPs that are not in the protein coding region do not alter the sequence of the protein, but may affect the expression of the gene by other means. For example, the SNP site located in the promoter region may affect the binding of the transcription factor, thereby affecting the transcription of the gene.
  • SNP is the third generation molecular marker following the first generation of molecularly labeled RFLP and the second generation of molecularly labeled microsatellite. It is commonly used in molecular marker technology based on DNA chip technology.
  • the HapMap program and the Thousand Genome Project have discovered a large number of SNP sites and representative tagSNP sites within a particular haplotype. So far, the dbSNP database has included more than 50 million confirmed SNP sites.
  • the principle of molecular marker technology based on DNA chip technology is to first design an allele-specific oligonucleotide probe for the tagSNP site in the genome-wide range, fix it on the chip, and then hybridize the DNA sample to be detected to the chip, and then The chip was scanned to obtain the genotype of the SNP site.
  • the most important application of molecular marker technology based on DNA chip technology is Genome-wide Association Studies (GWASs).
  • GWASs Genome-wide Association Studies
  • the usual research strategy of GWAS is: case-control strategy, which is to compare patients with diseases. And the genetic variation of the normal population (control) finds the susceptible site of the disease. Millions of genetically altered genotypes can be obtained from each person's DNA sample via the SNP chip.
  • a type of mutation (an allele) occurs more frequently in a patient, then the SNP is said to be associated with the disease.
  • These disease-related SNPs are used to label areas of the human genome that may affect the risk of disease.
  • the GWAS study targets the entire genome compared to methods that test only for genetic variation in one or more gene regions. Therefore, unlike studies for specific genomic regions, GWAS does not require prior selection of candidate genomic regions for the entire genome. GWAS is able to detect SNPs and other mutations in disease-associated susceptible sites, but it is not possible to determine specific pathogenic genes based solely on correlation.
  • Affymetrix is a pioneer in the gene chip industry. It developed the world's first gene chip as early as 1989.
  • the light-controlled in situ lithography of DNA microarrays is the highest density chip preparation technology available.
  • the hardware platform of the Affymetrix GeneChip chip system consists of a highly automated fluid workstation, a high-throughput chip scanner, and associated probe sequence description and annotation databases. Highly automated processing reduces manual time and increases data repeatability.
  • the Affymetrix chip uses in-situ lithography and rigorous process control to synthesize high-density gene chips that can synthesize more than 4 million probes per square centimeter of substrate.
  • the Affymetrix chip uses a unique PM-MM probe design that designs a pair of 25-mer probes for each reference sequence, one of which is a perfect match (PM) probe and the other is an error near the middle of the sequence. Site match (mismatch, MM) probe.
  • the detection signals of each pair of PM-MM probes are combined at the time of detection, which helps to distinguish the specific binding and non-specifically bound target fragments, thereby improving the sensitivity and specificity of the probe.
  • This PM-MM design has significant advantages in the detection of low abundance expression products in complex sequence background samples.
  • the use of multiple probes to detect SNPs effectively reduces the non-specificity of probe hybridization and obtains more robust data through appropriate algorithms.
  • the object of the present invention is to provide a DNA chip for SNPs in the non-coding region of the whole genome of the East Asian population, in particular, a SNP for the long-chain non-coding gene region and the miRNA gene region in the whole genome of the East Asian population. DNA chip.
  • the DNA chip of the SNPs for the non-coding region in the whole genome of the East Asian population provided by the present invention is a DNA chip immobilized with a specific probe; the specific probe is a probe for detecting 3568 SNPs in Table 1 ( The information of 3568 SNPs is shown in the first column and the second column of Table 1.
  • the specific probe may specifically consist of 4119 probes: a single-stranded DNA molecule represented by SEQ ID NO: 1 of the Sequence Listing to a single-stranded DNA molecule of SEQ ID NO: 4119 of the Sequence Listing.
  • long-chain non-coding gene regions is based on the non-coding RNA database independently developed by the applicant (Xie et al, NONCODEv4: exploring the world of long non-coding RNA genes. Nucleic Acids Res. 2014, 42: D98-D103.http: //www.noncode.org/).
  • the NONCODE database has been dedicated to the collection of non-coding RNAs of various species (except rRNA and tRNA) since about 10 years ago, and has been updated to the fourth version to date, including the latest 54072 long-chain non-coding genes from humans.
  • the inventors of the present invention have also collected the latest releases of the GENCODE program (Harrow et al., GENCODE: the reference human genome annotation for The ENCODE Project. Genome Research. 2012. 22: 1760-74. http://www.gencodegenes.org/). Human long-chain non-coding RNA data and Human lincRNA catalog data set (Cabili et al., Integrative annotation of human large intergenic noncoding RNAs reveals global properties and specific subclasses. 2011. Genes Dev 25: 1915-27). Based on the above data, a total of 26,977 long-chain non-coding gene regions were obtained.
  • the definition of the miRNA gene region is based on the 1877 miRNA precursor sequences of humans included in the NONCODE database. These sequences are also included in the miRBase database (Kozomara et al., miRBase: annotating high confidence microRNAs using deep sequencing data. Nucleic Acids Res. 2014.42: D68- D73).
  • SNP loci and genotypic data were obtained from the latest data published by the Thousand Human Genome Project, and only genotype data for common SNP loci in East Asian populations were considered. All SNP sites were grouped according to non-coding gene regions, then the r2 values between all SNP sites in each group were calculated, and a representative SNP site was selected, called tagSNP. A total of 3568 tag SNPs located in the non-coding gene region unique to the East Asian population were screened.
  • the information of 3568 tag SNP sites and the probe information for detecting the SNP sites are shown in Table 1.
  • Probes are single-stranded DNA molecules.
  • the probes for detecting each of the above SNP sites were immobilized on a substrate by Affymetrix, respectively, to obtain a genome-wide non-target for the East Asian population.
  • DNA chip of the SNPs of the coding region (Axiom array plate).
  • the DNA chips of the SNPs prepared for the non-coding regions in the whole genome of the East Asian population were tested using the DNA chips prepared in Example 2, and the specific methods are as follows:
  • step 2 Take about 200 ng of the genomic DNA obtained in step 1, using a kit (Axiom 2.0 Reagent Kit; Affymetrix product, article number 901758) and operating according to the kit instructions, and the whole genome-wide range for the East Asian population provided in Example 2. DNA chip hybridization of SNPs in non-coding regions, and then imaging detection on GeneTitan multichannel instrument (A/T is labeled with one color, G/C is labeled with another color), and the volunteers to be tested are based on each SNP site. Genotype.
  • the SNP chip currently used for GWAS mainly focuses on SNP sites related to protein-coding genes, including SNP sites located in the exon region and transcriptional regulatory region of the protein-coding gene, and there is no internationally targeted non-coding gene in the genome-wide range.
  • long-chain non-coding RNAs encode SNP chips, and the relationship between long-chain non-coding RNA and disease has been increasingly confirmed.
  • the present invention provides a DNA chip for SNPs of non-coding regions in the whole genome of East Asian population, which comprises screening from 26977 long-chain non-coding gene regions and 1877 miRNA gene regions. Of the 3568 SNPs, these SNPs are representative of SNPs in the transcribed regions of human non-coding genes.
  • the invention has great application value for detecting SNP sites in human long-chain non-coding gene regions, and has great application prospects for risk assessment of genetic diseases and personalized treatment.

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  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Wood Science & Technology (AREA)
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  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention concerne une puce à ADN des SNP dans une région non codante à l'intérieur de la plage du génome entier de la population d'Asie de l'Est, en particulier, une puce à ADN des SNP dans une longue région de gène non codante et une région de gène d'ARNmi à l'intérieur de la plage du génome entier de la population d'Asie de l'Est. La puce à ADN des SNP dans une région non codante à l'intérieur de la plage du génome entier de la population d'Asie de l'Est est une puce à ADN fixée avec une sonde spécifique ; la sonde spécifique est une sonde destinée à la détection de 3 568 SNP dans la Liste 1, et voir la première et la seconde colonne dans la Liste 1 pour plus d'information sur les 3 568 SNP. La puce présente une grande valeur d'application pour la détection de loci de SNP dans une longue région de gène non codante des êtres humains, et présente de grande perspective d'évaluation des risques et de traitement personnalisé de maladies héréditaires.
PCT/CN2016/070641 2016-01-12 2016-01-12 Puce à adn d'une région non codante des snp à l'intérieur de la plage du génome entier de la population d'asie de l'est WO2017120750A1 (fr)

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Application Number Priority Date Filing Date Title
PCT/CN2016/070641 WO2017120750A1 (fr) 2016-01-12 2016-01-12 Puce à adn d'une région non codante des snp à l'intérieur de la plage du génome entier de la population d'asie de l'est
CN201680000526.7A CN107018668B (zh) 2016-01-12 2016-01-12 一种针对东亚人群全基因组范围内的非编码区的SNPs的DNA芯片

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PCT/CN2016/070641 WO2017120750A1 (fr) 2016-01-12 2016-01-12 Puce à adn d'une région non codante des snp à l'intérieur de la plage du génome entier de la population d'asie de l'est

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CN110885888B (zh) * 2018-09-07 2022-04-29 中国科学院北京基因组研究所 用于推断亚洲不同地理区域人群的snp标记组合

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1438325A (zh) * 2003-03-10 2003-08-27 东南大学 一种基因组dna微阵列芯片及其制备和使用方法
WO2011074510A1 (fr) * 2009-12-14 2011-06-23 トヨタ自動車株式会社 Procédé de conception d'une sonde dans une puce à adn, et puce à adn munie d'une sonde conçue de cette manière
CN102121046A (zh) * 2009-06-25 2011-07-13 中国科学院北京基因组研究所 中国人群连锁分析snp标记集合及其使用方法与应用
WO2011083312A1 (fr) * 2010-01-08 2011-07-14 Oxford Gene Technology (Operations) Ltd Procédé combiné de cgh et d'hybridation allèle-spécifique

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CN101845501A (zh) * 2010-05-18 2010-09-29 孟涛 一种复杂疾病易感性综合遗传分析方法

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1438325A (zh) * 2003-03-10 2003-08-27 东南大学 一种基因组dna微阵列芯片及其制备和使用方法
CN102121046A (zh) * 2009-06-25 2011-07-13 中国科学院北京基因组研究所 中国人群连锁分析snp标记集合及其使用方法与应用
WO2011074510A1 (fr) * 2009-12-14 2011-06-23 トヨタ自動車株式会社 Procédé de conception d'une sonde dans une puce à adn, et puce à adn munie d'une sonde conçue de cette manière
WO2011083312A1 (fr) * 2010-01-08 2011-07-14 Oxford Gene Technology (Operations) Ltd Procédé combiné de cgh et d'hybridation allèle-spécifique

Non-Patent Citations (2)

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Title
CABILI, M.N. ET AL.: "Integrative annotation of human large intergenic noncoding RNAs reveals global properties and specific subclasses", GENES & DEVELOPMENT, vol. 25, 31 December 2011 (2011-12-31), pages 1915 - 1927, XP055400543 *
XIE, CHAOYONG ET AL.: "NONCODEv4: exploring the world of long non-coding RNA genes", NUCLEIC ACIDS RESEARCH, vol. 42, 26 November 2013 (2013-11-26), pages D98 - D103 *

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