EP1292707A2 - Oligonucleotides ou oligomeres de pna, et procede de detection parallele de l'etat de methylation d'un adn genomique - Google Patents

Oligonucleotides ou oligomeres de pna, et procede de detection parallele de l'etat de methylation d'un adn genomique

Info

Publication number
EP1292707A2
EP1292707A2 EP01927582A EP01927582A EP1292707A2 EP 1292707 A2 EP1292707 A2 EP 1292707A2 EP 01927582 A EP01927582 A EP 01927582A EP 01927582 A EP01927582 A EP 01927582A EP 1292707 A2 EP1292707 A2 EP 1292707A2
Authority
EP
European Patent Office
Prior art keywords
artificial sequence
dna
oligonucleotide
description
cgtacg
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP01927582A
Other languages
German (de)
English (en)
Inventor
Kurt Berlin
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Epigenomics AG
Original Assignee
Epigenomics AG
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Epigenomics AG filed Critical Epigenomics AG
Publication of EP1292707A2 publication Critical patent/EP1292707A2/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/82—Translation products from oncogenes
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • C07K14/4701—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals not used
    • C07K14/4702—Regulators; Modulating activity
    • C07K14/4703—Inhibitors; Suppressors
    • 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
    • C12Q1/6813—Hybridisation assays
    • C12Q1/6834—Enzymatic or biochemical coupling of nucleic acids to a solid phase
    • C12Q1/6837—Enzymatic or biochemical coupling of nucleic acids to a solid phase using probe arrays or probe chips
    • 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
    • C12Q1/6844—Nucleic acid amplification reactions
    • C12Q1/6853—Nucleic acid amplification reactions using modified primers or templates

Definitions

  • the second variant is based on partial chemical cleavage of total DNA, modeled on a Maxam-Gilbert sequencing reaction, ligation of adapters to the ends thus generated, amplification with generic primers and separation on a gel electrophoresis.
  • This method defined areas up to the size of less than a thousand base pairs can be examined.
  • the process is so complicated and unreliable that it is practically no longer used (Ward, C. et al., J. Biol. Chem. 265, 3030-3033).
  • the present invention describes a method and a set of oligonucleotides or PNA oligomers for the parallel detection of the methylation state : ⁇ ( ⁇ ) 90ivi99i ⁇ ( ⁇ ): ⁇ ( ⁇ ) 90ivi990 ⁇ ( ⁇ ) * ⁇ ( ⁇ ) 9 ⁇ vi99i ⁇ ( ⁇ )
  • a genomic DNA sample is chemically treated in such a way that at the 5'-position unmethylated cytosine bases are converted into uracil, thymine or another base which is unlike cytosine in terms of hybridization behavior.
  • the treatment of genomic DNA with bisulfite (bisulfite, disulfite) and subsequent alkaline hydrolysis, which leads to a conversion of unmethylated cytosine nucleobases into uracil, is preferably used for this purpose.
  • more than 26 different oligonucleotides are used simultaneously to produce a complex amplificate.
  • the products obtained after the amplification are separated by gel electrophoresis and the fragments which are smaller than 2000 base pairs or smaller than any limit below
  • the CpG dinucleotides contained in the amplified fragments are also examined completely or partially by hybridization of the fragments already provided with a detectable label in the amplification to a set of PNA oligomers which comprises at least two of the following sequences:
  • W one of the bases adenine (A) or thymine (T)
  • the first step is a genomic sequence using bisulfite
  • Figure 1 Here a high-density DNA chip is shown after hybridization.
  • the false color image is shown as it is generated after scanning. Contrary to the black and white illustration shown here, the scanner turns a colored one

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Zoology (AREA)
  • Molecular Biology (AREA)
  • Wood Science & Technology (AREA)
  • Engineering & Computer Science (AREA)
  • Genetics & Genomics (AREA)
  • General Health & Medical Sciences (AREA)
  • Biophysics (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biotechnology (AREA)
  • Medicinal Chemistry (AREA)
  • Analytical Chemistry (AREA)
  • Microbiology (AREA)
  • General Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Physics & Mathematics (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Toxicology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Oncology (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

L'invention concerne un procédé et un jeu d'oligonucléotides ou d'oligomères de PNA pour la détection parallèle de l'état de méthylation d'un ADN génomique. Selon l'invention, l'ADN est d'abord traité avec du bisulfites et cet ADN chimiquement modifié est ensuite fragmenté. Dans l'étape suivante, on procède à l'amplification de différents fragments avec des amorces synthétiques. Les produits de l'amplification sont hybridés avec des oligonucléotides d'une séquence connue et enfin détectés.
EP01927582A 2000-03-15 2001-03-15 Oligonucleotides ou oligomeres de pna, et procede de detection parallele de l'etat de methylation d'un adn genomique Withdrawn EP1292707A2 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
DE10013847A DE10013847A1 (de) 2000-03-15 2000-03-15 Oligonukleotide oder PNA-Oligomere und Verfahren zur parallelen Detektion des Methylierungszustandes genomischer DNA
DE10013847 2000-03-15
PCT/DE2001/001089 WO2001068910A2 (fr) 2000-03-15 2001-03-15 Oligonucleotides ou oligomeres de pna, et procede de detection parallele de l'etat de methylation d'un adn genomique

Publications (1)

Publication Number Publication Date
EP1292707A2 true EP1292707A2 (fr) 2003-03-19

Family

ID=7635678

Family Applications (1)

Application Number Title Priority Date Filing Date
EP01927582A Withdrawn EP1292707A2 (fr) 2000-03-15 2001-03-15 Oligonucleotides ou oligomeres de pna, et procede de detection parallele de l'etat de methylation d'un adn genomique

Country Status (5)

Country Link
US (1) US20050202420A1 (fr)
EP (1) EP1292707A2 (fr)
AU (1) AU2001254601A1 (fr)
DE (1) DE10013847A1 (fr)
WO (1) WO2001068910A2 (fr)

Families Citing this family (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2003144172A (ja) * 2001-11-16 2003-05-20 Nisshinbo Ind Inc メチル化検出用オリゴヌクレオチド固定化基板
US7932027B2 (en) 2005-02-16 2011-04-26 Epigenomics Ag Method for determining the methylation pattern of a polynucleic acid
WO2006088978A1 (fr) 2005-02-16 2006-08-24 Epigenomics, Inc. Procede de determination du modele de methylation d'un acide polynucleique
JP5133238B2 (ja) 2005-04-15 2013-01-30 エピゲノミックス アクチェンゲゼルシャフト 遠隔サンプル由来のｄｎａ断片を提供する方法
US7820385B2 (en) 2006-03-22 2010-10-26 The United States Of America As Represented By The Department Of Health And Human Services, Centers For Disease Control And Prevention Method for retaining methylation pattern in globally amplified DNA
US8084734B2 (en) * 2006-05-26 2011-12-27 The George Washington University Laser desorption ionization and peptide sequencing on laser induced silicon microcolumn arrays
US8110796B2 (en) 2009-01-17 2012-02-07 The George Washington University Nanophotonic production, modulation and switching of ions by silicon microcolumn arrays
US9490113B2 (en) * 2009-04-07 2016-11-08 The George Washington University Tailored nanopost arrays (NAPA) for laser desorption ionization in mass spectrometry

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1991000343A2 (fr) * 1989-06-29 1991-01-10 Gist-Brocades N.V. Enzyme mutante ayant une stabilite reduite dans des conditions d'application industrielle
US6017704A (en) * 1996-06-03 2000-01-25 The Johns Hopkins University School Of Medicine Method of detection of methylated nucleic acid using agents which modify unmethylated cytosine and distinguishing modified methylated and non-methylated nucleic acids
DE19754482A1 (de) * 1997-11-27 1999-07-01 Epigenomics Gmbh Verfahren zur Herstellung komplexer DNA-Methylierungs-Fingerabdrücke
WO1999055914A1 (fr) * 1998-04-29 1999-11-04 Trustees Of Boston University Compositions et procedes relatifs aux boucles pd
WO2000001816A1 (fr) * 1998-07-02 2000-01-13 Imperial Cancer Research Technology Limited GENE SUPPRESSEUR DE TUMEUR DBCCR1 SITUE DANS 9q32-33
US6107091A (en) * 1998-12-03 2000-08-22 Isis Pharmaceuticals Inc. Antisense inhibition of G-alpha-16 expression
US6136603A (en) * 1999-03-26 2000-10-24 Isis Pharmaceuticals Inc. Antisense modulation of interleukin-5 signal transduction

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
None *
See also references of WO0168910A3 *

Also Published As

Publication number Publication date
US20050202420A1 (en) 2005-09-15
WO2001068910A3 (fr) 2003-01-03
AU2001254601A1 (en) 2001-09-24
WO2001068910A2 (fr) 2001-09-20
DE10013847A1 (de) 2001-09-27

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