Automated system for polynucleotide synthesis and purification

• US 5,935,527 A
• Filed: 04/29/1993
• Issued: 08/10/1999
• Est. Priority Date: 12/21/1988
• Status: Expired due to Term

• Current Assignee: Applied Biosystems LLC (Thermo Fisher Scientific Incorporated)
• Original Assignee: The Perkin-Elmer Corporation (Thermo Fisher Scientific Incorporated)
• Inventors: McCollum, Christie D., Zon, Gerald, Andrus, William A.
• Primary Examiner(s): TRAN, HIEN THI
• Application Number:
  US08/055,464
• Time in Patent Office: 2,294 Days
• Field of Search: 422/131, 422/129, 536/27-29, 536/25.31, 935/88, 435/6
• US Class Current: 422/131
• 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 2219/00286 Reactor vessels with top and bottom openings
• CPC Class Codes: B01J 2219/00351 Means for dispensing and evacuation of reagents
• CPC Class Codes: B01J 2219/00389 Feeding through valves
• CPC Class Codes: B01J 2219/00454 by chemical cleavage from the solid support
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/0059 Sequential processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/0068 Means for controlling the apparatus of the process
• CPC Class Codes: B01J 2219/00686 Automatic
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C12N 15/1006 by means of a solid support carrier, e.g. particles, polymers
• CPC Class Codes: C40B 40/06 Libraries containing nucleotides or polynucleotides, 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: C40B 60/14 for creating libraries

Abstract

A method and system for polynucleotide synthesis are provided which employ solid phase synthesis on a nonswellable porous polystyrene support by phosphoramidite or hydrogen phosphonate chemistries. The polystyrene support gives rise to fewer tritylated failure sequences caused by chain growth from extraneous support sites, and allows lower amounts of monomer reactants to be used to achieve equal or better coupling efficiencies as those achieveable with CPG. The method and system also employ nucleoside intermediates whose exocyclic amines are protected by base-labile groups which permit simultaneous cleavage and deprotection of the completed polynucleotide chain in the presence of the solid phase support. This latter feature allows practical automation of both the synthesis and purification of polynucleotides.

Claims

1. A chamber for solid phase polynucleotide synthesis, the chamber comprising:

a chamber, and disposed in said chamber

(i) a nonswellable porous polystyrene support capable of remaining substantially mechanically rigid during polynucleotide synthesis and having a 5'"'"'-tritylated protected nucleoside attached covalently thereto, for allowing synthesis of a polynucleotide on the support, and

(ii) a hydrophobic adsorbent capable of non-covalently adsorbing a tritylated polynucleotide, for purifying the polynucleotide after synthesis of the polynucleotide on said nonswellable porous polystyrene support.

2. The chamber of claim 1, wherein said hydrophobic adsorbent is a nonswellable porous polystyrene adsorbent.