WO2020054907A1 - Oligonucléotides pour amplification de séquence cible et procédé d'amplification de séquence cible l'utilisant - Google Patents

Oligonucléotides pour amplification de séquence cible et procédé d'amplification de séquence cible l'utilisant Download PDF

Info

Publication number
WO2020054907A1
WO2020054907A1 PCT/KR2018/013318 KR2018013318W WO2020054907A1 WO 2020054907 A1 WO2020054907 A1 WO 2020054907A1 KR 2018013318 W KR2018013318 W KR 2018013318W WO 2020054907 A1 WO2020054907 A1 WO 2020054907A1
Authority
WO
WIPO (PCT)
Prior art keywords
amplification
oligonucleotide
target dna
dna sequence
sequence
Prior art date
Application number
PCT/KR2018/013318
Other languages
English (en)
Korean (ko)
Inventor
유석종
Original Assignee
한국과학기술정보연구원
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 한국과학기술정보연구원 filed Critical 한국과학기술정보연구원
Publication of WO2020054907A1 publication Critical patent/WO2020054907A1/fr

Links

Images

Classifications

    • 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
    • C12Q1/686—Polymerase chain reaction [PCR]
    • 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
    • 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
    • C12Q2521/00—Reaction characterised by the enzymatic activity
    • C12Q2521/30—Phosphoric diester hydrolysing, i.e. nuclease
    • C12Q2521/301—Endonuclease
    • 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
    • C12Q2527/00—Reactions demanding special reaction conditions
    • C12Q2527/101—Temperature

Definitions

  • the oligonucleotide for DNA amplification according to the present invention shows the amplification process using the oligonucleotide for DNA amplification according to the present invention.
  • the enzyme capable of operating Crispr-cas9 as the Cas 9 enzyme the oligonucleotide for DNA amplification according to the present invention hybridizes to the target DNA sequence, and Cas 9 (Catalytically Cas 9)
  • Cas 9 Catalytically Cas 9
  • the oligonucleotide for DNA amplification is characterized in that it is 5 'phosphorylated.
  • the oligonucleotide for DNA amplification is characterized in that it is a DNA sequence.
  • Oligonucleotides for amplification of a target DNA sequence can be designed in a variety of ways obvious to those skilled in the art, and the mutation of a gene in a gene to be amplified during gene amplification using multiple sequence alignment results and Shannon entropy It is possible to design by selecting the fewest areas.
  • first column entropy' of the present invention means a value obtained by calculating the degree of mutation in each column of a multi-sequence aligned file of genetic data using Shannon entropy. do.
  • the first column entropy value in the second step is calculated according to Equation 1 below.
  • the length is changed to a length of 15 to 25 bp while moving the starting position of the primer at 1 bp intervals from the first nucleotide sequence to the last nucleotide sequence, that is, all sequences of the multi-sequence genetic data sequence in the third step.
  • a combination of the forward primers and reverse primers was generated.
  • the weight ( ⁇ w ) of the gradient penalty may be defined by the user, and has a value of 0 or more, and the initial value is set to 1.
  • the primer entropy value in the fifth step is the following equation using the second column entropy Q (i) according to Equation 3 above. It is characterized by calculating according to 4.
  • Wl , W g , W t , W h , W d, and W e in Equation 5 above are weights for primer length,% gc, Tm, hairpin formation, dimer formation, and primer entropy, respectively.
  • W l , W g , W t , W h , W d and W e can be defined by the user, have a value of 0 or more, and the initial value is set to 1.
  • ⁇ f and ⁇ r are primer entropy for the forward and reverse primers calculated in Equation 4 above)
  • a primer set based on a reference value set by an experimenter by reflecting any one or more of a primer length, GC fraction, hairpin score, dimer score, and Tm value.
  • the primer length may be adopted in the range of 10-50 bp, preferably in the range of 10-40 bp, and most preferably in the range of 10-25 bp.
  • the Cas9 enzyme (Catalytically Cas 9) to which the oligonucleotide for DNA amplification reacts is characterized by loss of endonuclease function.
  • the method for amplifying a target DNA sequence according to the present invention can be PCR, LAMP, NGS.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Analytical Chemistry (AREA)
  • Biophysics (AREA)
  • Immunology (AREA)
  • Microbiology (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Physics & Mathematics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • 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 des oligonucléotides pour une amplification de séquence cible et un procédé d'amplification de séquence cible l'utilisant et, plus spécifiquement, des oligonucléotides, qui comprennent crispr-cas9 ou un système correspondant à celui-ci de façon à détecter, par l'intermédiaire d'une réaction avec une enzyme Cas 9, un emplacement de séquence cible auquel les oligonucléotides peuvent se lier, et sont liés à la séquence cible à température ambiante sans dispositifs spéciaux de façon à être amplifiés ; et un procédé d'amplification de séquence cible l'utilisant.
PCT/KR2018/013318 2018-09-12 2018-11-05 Oligonucléotides pour amplification de séquence cible et procédé d'amplification de séquence cible l'utilisant WO2020054907A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
KR10-2018-0108615 2018-09-12
KR1020180108615A KR102200109B1 (ko) 2018-09-12 2018-09-12 표적 서열 증폭용 올리고 뉴클레오타이드 및 이를 이용한 표적 서열의 증폭 방법

Publications (1)

Publication Number Publication Date
WO2020054907A1 true WO2020054907A1 (fr) 2020-03-19

Family

ID=69777215

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/KR2018/013318 WO2020054907A1 (fr) 2018-09-12 2018-11-05 Oligonucléotides pour amplification de séquence cible et procédé d'amplification de séquence cible l'utilisant

Country Status (2)

Country Link
KR (1) KR102200109B1 (fr)
WO (1) WO2020054907A1 (fr)

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20150138074A (ko) * 2014-05-28 2015-12-09 주식회사 툴젠 표적 특이적 뉴클레아제를 이용한 표적 dna의 민감한 검출 방법
WO2017100343A1 (fr) * 2015-12-07 2017-06-15 Arc Bio, Llc Procédés et compositions pour la fabrication et l'utilisation d'acides nucléiques de guidage

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9850525B2 (en) 2014-01-29 2017-12-26 Agilent Technologies, Inc. CAS9-based isothermal method of detection of specific DNA sequence
CN109790537A (zh) 2016-09-29 2019-05-21 富士胶片株式会社 供多重pcr的引物的设计方法

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20150138074A (ko) * 2014-05-28 2015-12-09 주식회사 툴젠 표적 특이적 뉴클레아제를 이용한 표적 dna의 민감한 검출 방법
WO2017100343A1 (fr) * 2015-12-07 2017-06-15 Arc Bio, Llc Procédés et compositions pour la fabrication et l'utilisation d'acides nucléiques de guidage

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
GAO, F.: "DNA-guided genome editing using the Natronobacterium gregoryi Argonaute", NATURE BIOTECHNOLOGY, vol. 34, no. 7, 2 May 2016 (2016-05-02), pages 768 - 773, XP055518128 *
VARSHNEY, G. K.: "DNA-guided genome editing using structure-guided endonucleases", GENOME BIOLOGY, vol. 17, no. 1, 187, 15 September 2016 (2016-09-15), pages 1 - 4, XP055419357 *
YIN, H.: "Partial DNA-guided Cas9 enables genome editing with reduced off-target activity", NATURE CHEMICAL BIOLOGY, vol. 14, no. 3, 29 January 2018 (2018-01-29), pages 311 - 316, XP055515605, DOI: 10.1038/nchembio.2559 *

Also Published As

Publication number Publication date
KR20200030166A (ko) 2020-03-20
KR102200109B1 (ko) 2021-01-08

Similar Documents

Publication Publication Date Title
US11692213B2 (en) Compositions and methods for targeted depletion, enrichment, and partitioning of nucleic acids using CRISPR/Cas system proteins
US20210277389A1 (en) Methods and Compositions for the Single Tube Preparation of Sequencing Libraries Using Cas9
Matic et al. Genetic barriers among bacteria
KR101293713B1 (ko) 시토신의 화학적 변형에 의한 미생물 핵산의 단순화 방법
US20190194648A1 (en) Construction method for serial sequencing libraries of rad tags
US20050064406A1 (en) Methods for high throughput genome analysis using restriction site tagged microarrays
KR900006520A (ko) 결실 탐지를 위한 게놈성 dna다중 증폭방법
JP2005504508A5 (fr)
CA2283651A1 (fr) Extraction et utilisation d'alleles vntr
JP2009538603A (ja) 微生物の検出および分析に使用するための微生物改変核酸
JP7436493B2 (ja) 次世代シーケンスにおいて低サンプルインプットを扱うための正規化対照
Glick et al. Medical biotechnology
Książczyk et al. Genome-dependent chromosome dynamics in three successive generations of the allotetraploid Festuca pratensis× Lolium perenne hybrid
EP3872171A1 (fr) Détection d'arn et édition dépendant de la transcription à l'aide d'arncr transactivateurs reprogrammés
WO2020054907A1 (fr) Oligonucléotides pour amplification de séquence cible et procédé d'amplification de séquence cible l'utilisant
CN104388456A (zh) 一种同时表达两条sgRNA的载体的构建方法
Groathouse et al. Isothermal amplification and molecular typing of the obligate intracellular pathogen Mycobacterium leprae isolated from tissues of unknown origins
US8470537B2 (en) Sequential addition of short DNA oligos in DNA-polymerase-based synthesis reactions
Khalil Different types of PCR
JP2004516801A (ja) エシェリキア・コリ菌株ｄｓｍ６６０１の同定法
Tang et al. Single transcript unit CRISPR 2.0 systems for genome editing in rice
CA2393874C (fr) Procede permettant d'isoler de maniere selective un acide nucleique
US20030044785A1 (en) Method for identifying and genotyping microorganisms
Buckingham Fundamentals of Nucleic Acid Biochemistry: An Overview
Bartoli et al. Rapid protocol for template preparation of large numbers of clones

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 18933610

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 18933610

Country of ref document: EP

Kind code of ref document: A1