DK2398915T3 - Syntese af sekvensverificerede nukleinsyrer - Google Patents
Syntese af sekvensverificerede nukleinsyrer Download PDFInfo
- Publication number
- DK2398915T3 DK2398915T3 DK10705856.2T DK10705856T DK2398915T3 DK 2398915 T3 DK2398915 T3 DK 2398915T3 DK 10705856 T DK10705856 T DK 10705856T DK 2398915 T3 DK2398915 T3 DK 2398915T3
- Authority
- DK
- Denmark
- Prior art keywords
- nucleic acid
- sequence
- sequencing
- dna
- fragments
- Prior art date
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
- C12N15/1034—Isolating an individual clone by screening libraries
- C12N15/1093—General methods of preparing gene libraries, not provided for in other subgroups
-
- 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
- C12Q1/6869—Methods for sequencing
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Microbiology (AREA)
- Biomedical Technology (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Plant Pathology (AREA)
- Crystallography & Structural Chemistry (AREA)
- Bioinformatics & Computational Biology (AREA)
- Analytical Chemistry (AREA)
- Immunology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Claims (14)
1. Fremgangsmåde til automatisk indvinding af sekvensverificerede nukleinsyrer fra en plade, en bærer og/eller et substrat, hvilken fremgangsmåde omfatter følgende trin: (1) at tilvejebringe en blanding af nukleinsyrer, (2) at omdanne nukleinsyreblandingen til et klonalt bibliotek, som er til stede på pladen, bæreren og/eller substratet, ved at individualisere nukleinsyrerne i blandingen fra (1), (3) at sekventere det klonale bibliotek parallelt, hvor, som resultat deraf, sekvensen og placeringen er kendt for hvert enkelt molekyle i det klonale bibliotek, og (4) at indvinde de sekvensverificerede nukleinsyremolekyler, hvor indvindingen omfatter computerstøttet lokalisering af de sekvensverificerede nukleinsyremolekyler på et mikroskopibillede af pladen, bæreren og/eller substratet ved hjælp af referencepunkter, og hvor referencepunkternes placeringer identificeres på mikroskopibilledet af bæreren ved hjælp af det klonale DNA med kendt sekvens.
2. Fremgangsmåde ifølge krav 1, hvor bæreren er valgt blandt objektglas, geler, polymerer, kapillarrør, mikrofluidbærere, membraner, porøse bærere, plast, silicium, ordnede eller kaotiske porer, svampestrukturer, terninger, 2D og 3D, emulsioner, dendrimerer, perler, partikler, resiner, metaller, nanopartikler og nanostrukturer eller kombinationer deraf.
3. Fremgangsmåde ifølge et hvilket som helst af kravene 1-2, hvor bæreren omfatter perler.
4. Fremgangsmåde ifølge krav 3, hvor den computerstøttede lokalisering gør brug af referencepunkter ved at anvende farvede eller fluorescerende perler, der bærer det klonale DNA med kendt sekvens, som kan lokaliseres i sekventerings-apparatet og på mikroskopibilledet.
5. Fremgangsmåde ifølge et hvilket som helst af kravene 1-4, hvor indvindingen i trin (4) omfatter afkopiering eller kopiering og amplifikation af de bærer-bundne nukleinsyremolekyler.
6. Fremgangsmåde ifølge et hvilket som helst af kravene 1-5, hvor indvindingen omfatter laserudslyngning, laserindfangning, brug af laserpincetter og/eller laserskæring.
7. Fremgangsmåde ifølge et hvilket som helst af kravene 1-6, hvor bæreren tilvejebringes som eller i en mikrotiterplade, en nanotiterplade og/eller en picoti-terplade.
8. Fremgangsmåde ifølge et hvilket som helst af kravene 1 -7, hvor trin (3) omfatter Sanger-sekventering, næstegenerationssekventering, sekventering-ved-syntese, sekventering-ved-ligering, enkeltmolekyle-sekventering eller nanopore-sekventering.
9. Fremgangsmåde ifølge et hvilket som helst af kravene 1 -8, hvor trin (2) omfatter emulsions-PCR, broamplifikation, MegaPlex-PCR, frembringelse af polonier og/eller rullecirkel-amplifikation (rolling circle amplification), hvor emulsions-PCR foretrækkes.
10. Fremgangsmåde ifølge et hvilket som helst af kravene 1-9, som endvidere omfatter trinet (5) til dannelse af en eller flere pools af sekvensverificerede nukleinsyremolekyler.
11. Apparat til fremstilling af sekvensverificerede nukleinsyrefragmenter, hvilket apparat indbefatter (a) en enhed/et middel til at generere en klonal repræsentation af en nukleinsy-reblanding på en plade, en bærer og/eller et substrat, hvor den klonale repræsentation af en nukleinsyreblanding fortrinsvis omfatter monokloniserede nukleinsyrer opnået fra blandingen, (b) en enhed/et middel til at sekventere den klonale repræsentation af nukleinsy-reblandingen og generere sekvensinformationen og informationen om den fysiske placering for hvert af nukleinsyrefragmenterne, (c) en enhed/et middel til at forbinde sekvensinformationen med den fysiske position eller placering af nukleinsyrefragmenterne, hvor enheden/midlet omfatter et middel til at identificere den fysiske position eller placering af nukleinsyrefragmenterne ved hjælp af referencepunkter på et mikroskopibillede af pladen, bæreren og/eller substratet, (d) en enhed/et middel til at frigøre de sekvensverificerede nukleinsyrefragmenter fra bæreren på grundlag af den tilvejebragte sekvens og placeringsinformation og (e) en enhed/et middel til at indvinde de frigjorte sekvensverificerede nukleinsyrefragmenter fra bæreren.
12. Apparat til automatisk indvinding af sekvensverificerede nukleinsyrefragmenter, hvilket apparat indbefatter (a) en enhed/et middel til at registrere et mikroskopibillede af en bærer, som omfatter perler opnået fra parallel sekventering af et klonalt bibliotek af en nuklein-syreblanding, (b) en enhed/et middel til at tilvejebringe placerings- og sekvensinformationen for hvert af de klonale nukleinsyrefragmenter på bæreren fra (a), (c) en enhed/et middel til at forbinde sekvensinformationen med den fysiske position eller placering af nukleinsyrefragmenterne, hvor enheden/midlet omfatter et middel til at identificere den fysiske position eller placering af nukleinsyrefragmenterne ved hjælp af referencepunkter på mikroskopibilledet af bæreren fra (a), (d) en enhed/et middel til at indvinde nukleinsyrefragmenterne fra bæreren på grundlag af den i trin (c) tilvejebragte sekvens og placeringsinformation og (e) en enhed/et middel til at frigøre de sekvensverificerede nukleinsyrefragmenter fra perlerne.
13. Apparat ifølge krav 11 eller 12, hvor referencepunkterne omfatter farvede eller fluorescerende perler, der bærer veldefinerede DNA-sekvenser, som kan lokaliseres i sekventeringsapparatet og på mikroskopibilledet.
14. Apparat ifølge et hvilket som helst af kravene 11-13, som indeholder yderligere en enhed/et middel til at reformatere de indvundne sekvensverificerede nu-kleinsyrefragmenter.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US15409109P | 2009-02-20 | 2009-02-20 | |
| PCT/EP2010/052140 WO2010094772A1 (en) | 2009-02-20 | 2010-02-19 | Synthesis of sequence-verified nucleic acids |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| DK2398915T3 true DK2398915T3 (da) | 2016-12-12 |
Family
ID=42236973
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| DK10705856.2T DK2398915T3 (da) | 2009-02-20 | 2010-02-19 | Syntese af sekvensverificerede nukleinsyrer |
Country Status (4)
| Country | Link |
|---|---|
| US (2) | US20100216648A1 (da) |
| EP (1) | EP2398915B1 (da) |
| DK (1) | DK2398915T3 (da) |
| WO (1) | WO2010094772A1 (da) |
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-
2010
- 2010-02-19 US US12/708,783 patent/US20100216648A1/en not_active Abandoned
- 2010-02-19 WO PCT/EP2010/052140 patent/WO2010094772A1/en active Application Filing
- 2010-02-19 EP EP10705856.2A patent/EP2398915B1/en active Active
- 2010-02-19 DK DK10705856.2T patent/DK2398915T3/da active
-
2017
- 2017-03-21 US US15/465,532 patent/US10876110B2/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| WO2010094772A1 (en) | 2010-08-26 |
| US10876110B2 (en) | 2020-12-29 |
| US20100216648A1 (en) | 2010-08-26 |
| EP2398915A1 (en) | 2011-12-28 |
| EP2398915B1 (en) | 2016-08-24 |
| US20170267999A1 (en) | 2017-09-21 |
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