Method for producing polymers

• US 20030198948A1
• Filed: 06/06/2003
• Published: 10/23/2003
• Est. Priority Date: 02/19/1999
• Status: Active Grant

• Current Assignee: Codex DNA, Inc.
• Original Assignee: Febit Holding GmbH (FEBIT)
• Inventors: Muller, Manfred, Stahler, Cord F., Stahler, Peer F.
• Granted Patent: US 7,790,369 B2
• Time in Patent Office: Days
• US Class Current: 435/5
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 19/0093 Microreactors, e.g. miniaturised or microfabricated reactors laboratory containers with capillary fluid transport in microfabricated channels or chambers B01L3/5027
• CPC Class Codes: B01J 2219/00317 Microwell devices, i.e. having large numbers of wells
• CPC Class Codes: B01J 2219/00432 Photolithographic masks
• CPC Class Codes: B01J 2219/00436 Maskless processes
• CPC Class Codes: B01J 2219/00439 using micromirror arrays
• CPC Class Codes: B01J 2219/00441 using lasers
• CPC Class Codes: B01J 2219/00448 using microlens arrays
• CPC Class Codes: B01J 2219/00479 Means for mixing reactants or products in the reaction vessels
• CPC Class Codes: B01J 2219/00497 Features relating to the solid phase supports
• CPC Class Codes: B01J 2219/00511 Walls of reactor vessels
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/0059 Sequential processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00603 Making arrays on substantially continuous surfaces
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00648 : by the use of solid beads
• CPC Class Codes: B01J 2219/00659 : Two-dimensional arrays
• CPC Class Codes: B01J 2219/00675 : In-situ synthesis on the substrate
• CPC Class Codes: B01J 2219/00689 : using computers
• CPC Class Codes: B01J 2219/00702 : Processes involving means for analysing and characterising the products
• CPC Class Codes: B01J 2219/00704 : integrated with the reactor apparatus
• CPC Class Codes: B01J 2219/00711 : Light-directed synthesis
• CPC Class Codes: B01J 2219/00722 : Nucleotides
• CPC Class Codes: B01J 2219/00725 : Peptides
• CPC Class Codes: B01L 2300/0654 : Lenses; Optical fibres
• CPC Class Codes: B01L 2300/069 : Absorbents; Gels to retain a fluid
• CPC Class Codes: B01L 2300/0864 : comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 3/502707 : characterised by the manufacture of the container or its components by shaping or joining plastic parts B29C59/00, B29C65/00; by laminating B32B37/00; manufacture of microstructural devices in general B81C
• CPC Class Codes: B01L 3/5085 : for multiple samples, e.g. microtitration plates
• CPC Class Codes: B82Y 30/00 : Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C12N 15/10 : Processes for the isolation, preparation or purification of DNA or RNA chemical preparation of DNA or RNA C07H21/00; preparation of non-structural polynucleotides from microorganisms or with enzymes C12P19/34
• CPC Class Codes: C12N 15/66 : General methods for inserting a gene into a vector to form a recombinant vector using cleavage and ligation; Use of non-functional linkers or adaptors, e.g. linkers containing the sequence for a restriction endonuclease
• CPC Class Codes: C12P 19/34 : Polynucleotides, e.g. nucleic acids, oligoribonucleotides
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 40/10 : Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 50/14 : Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
• CPC Class Codes: G03F 7/70216 : Mask projection systems
• CPC Class Codes: G16B 30/00 : ICT specially adapted for sequence analysis involving nucleotides or amino acids
• CPC Class Codes: G16B 50/00 : ICT programming tools or database systems specially adapted for bioinformatics

Abstract

The invention relates to a method for producing polymers, in particular synthetic nucleic acid double strands of optional sequence, comprising the steps:

(a) provision of a support having a surface area which contains a plurality of individual reaction areas,

(b) location-resolved synthesis of nucleic acid fragments having in each case different base sequences in several of the individual reaction areas, and

(c) detachment of the nucleic acid fragments from individual reaction areas.

Claims

1. Method for synthesizing polymers,

characterized in that

a plurality of oligomeric building blocks is synthesized on a support by parallel synthesis steps, is detached from the support and is brought into contact with one another to synthesize the polymer.

2. Method according to claim 1,

characterized in that

double-stranded nucleic acid polymers of at least 300 bp, in particular at least 1000 bp in length are synthesized.

3. Method according to claim 1,

characterized in that

nucleic acid polymers selected from the group consisting of genes, gene clusters, chromosomes, viral and bacterial genomes or sections thereof are synthesized.

4. Method according to claim 1,

characterized in that

the oligomeric building blocks are from 5 to 150, preferably 5 to 30 monomer units in length.

5. Method according to claim 1,

characterized in that

in successive steps in each case partially complementary oligonucleotide building blocks are detached from the support and are brought into contact with one another or with the polymer intermediate under hybridization conditions.

6. Method according to claim 1 for producing synthetic nucleic acid double strands of optional sequence, comprising the steps:

(a) provision of a support having a surface area which contains a plurality of individual reaction areas,

(b) location-resolved synthesis of nucleic acid fragments having in each case different base sequences in several of the individual reaction areas, and

(c) detachment of the nucleic acid fragments from individual reaction areas.

7. Method according to claim 6,

characterized in that

the base sequences of the nucleic acid fragments synthesized in individual reaction areas are chosen such that they can assemble to form a nucleic acid double strand hybrid.

8. Method according to claim 6,

characterized in that

the nucleic acid fragments according to step (c) are detached in one or more steps under conditions such that a plurality of the detached nucleic acid fragments assemble to form a nucleic acid double strand hybrid.

9. Method according to claim 8,

characterized in that

several nucleic acid fragments which form one strand of the nucleic acid double strand hybrid are linked covalently to one another.

10. Method according to claim 9,

characterized in that

the covalent linking includes treatment with ligase or/and filling in gaps in the strands using DNA polymerase.

11. Method according to claim 6,

characterized in that

the sequence comprises at one or more positions recognition sequences for specific interaction with molecules such as proteins, nucleic acids, peptides, pharmaceuticals, saccharides, lipids, hormones, or/and organic compounds.

12. Method according to claim 6,

characterized in that

the sequence of the nucleic acid double strands is a naturally occurring sequence, a not naturally occurring sequence or a combination of these two.

13. Method according to claim 6,

characterized in that

the sequence is taken from a database, a sequencing experiment or a device for the integrated synthesis and analysis of polymers.

14. Method according to claim 1,

characterized in that

the oligomeric building blocks are synthesized by location- or/and time-resolved illumination by means of a programmable light source matrix.

15. Method according to claim 1,

characterized in that

a location- or/and time-resolved synthesis of the oligomeric building blocks takes place in a microfluidic reaction support having one or more fluidic reaction compartments and one or more reaction areas within a fluidic reaction compartment.

16. Method according to claim 1,

characterized in that

the synthesis building blocks contain nucleotides occurring in nature, modified nucleotides or mixtures thereof.

17. Method according to claim 1,

characterized in that

modified synthesis building blocks are used for labeling and subsequent detection of the assembled nucleic acid double strands.

18. Method according to claim 17,

characterized in that

the labeling groups used are molecules which are to be detected in a light-dependent manner.

19. Use of a nucleic acid double strand of optional sequence, produced according to the method according to claim 1, for therapeutic or pharmacological purposes.

20. Use of a nucleic acid double strand of optional sequence, produced according to the method according to claim 1, for diagnostic purposes.

21. Use according to claim 19, comprising direct application to the intended purpose.

22. Use according to claim 19, comprising a conversion in effector cells.

23. Use of a nucleic acid double strand of optional sequence, produced according to the method according to claim 1, where said nucleic acid double strand is stabilized, condensed or/and topologically manipulated during or following the combination and assembly in stages.

24. Use according to claim 23, where stabilization, condensation or/and topological manipulation is carried out by functional molecules such as histones or topoisomerases.

25. Use of a nucleic acid of claim 1 as a propagatable cloning vector, where the propagatable cloning vector may serve for transcription, expression of the transcribed sequence, and, where appropriate, production of expressed gene products in suitable target cells.