Biopolymer synthesis system and method

Abstract

The present invention provides improved automated systems and methods for synthesis of biopolymers including DNA and RNA. The automated systems and methods represent a number of improvements over existing systems for multiplex synthesis of biopolymers in a combinatorial fashion.

1 . A system for synthesizing a polymer, the system comprising:
 a reagent supply system;   a first manifold coupled to the reagent supply system;   a plurality of reaction vessels coupled to the first manifold, wherein each reaction vessel of the plurality of reaction vessels comprises a first porous membrane, a second porous membrane, and a region disposed between the first porous membrane and second porous membrane for containing a solid support medium;   a reagent control system coupled to the plurality of vessels, wherein the reagent control system is configured to selectively deliver reagent originating from the reagent supply system to a first subset of the plurality of reaction vessels; and   an outlet port coupled to the first manifold;   wherein the system is configured such that reagent from the reagent supply system passes through the system to the outlet port without contacting the ambient environment.   
     
     
         2 . The system of  claim 1 , further comprising a wash system coupled to the plurality of reaction vessels, wherein the wash system is configured to deliver wash fluid through each of the plurality of reaction vessels to exit from the outlet port. 
     
     
         3 . The system of  claim 2 , wherein wash fluid from the wash system enters into the reaction vessel through the second porous membrane and the first reagent initially enters into the reaction vessel through the first porous membrane. 
     
     
         4 . The system of  claim 1 , wherein each reaction vessel of the plurality of reaction vessels is orientated such that the first porous membrane is disposed below the second porous membrane. 
     
     
         5 . The system of  claim 1 , further comprising a plurality of branched connectors, wherein each branched connector couples a reaction vessel of the plurality of reaction vessels to the wash system and reagent control system. 
     
     
         6 . The system of  claim 1 , further comprising a second manifold that includes the branched connector. 
     
     
         7 . The system of  claim 1 , wherein the wash system further comprises a compressed inert gas, and wherein the wash system is configured to cause the flow of inert gas through each of the plurality of reaction vessels to exit from the outlet port. 
     
     
         8 . The system of  claim 1 , wherein the system is configured such that, in operation, the system carries out a reagent delivery step, a reaction step, and a reagent wash step. 
     
     
         9 . The system of  claim 8 , wherein the reagent delivery step comprises:
 flowing reagent from the reagent supply system through the first manifold; and   drawing the first reagent through the first porous membrane of the subset of the plurality of reaction vessels into the region for containing the solid support medium.   
     
     
         10 . The system of  claim 8 , wherein the reaction step comprises repeatedly displacing the first reagent across the subset of reaction vessels. 
     
     
         11 . The system of  claim 8 , wherein the reagent wash step comprises delivering a wash fluid through the second porous membrane into the plurality of reaction vessels, through the first manifold to exit the outlet port. 
     
     
         12 . The system of  claim 1 , wherein each reagent of the plurality of reagents is pressurized within a container. 
     
     
         13 . The system of  claim 1 , wherein the reagent control system is substantially devoid of wash fluid or first reagent throughout operation of the system. 
     
     
         14 . The system of  claim 1 , further comprising a reaction plate that contains and/or defines the plurality of reaction vessels. 
     
     
         15 . The system of  claim 14 , wherein the reaction plate is temperature-controlled. 
     
     
         16 . The system of  claim 14 , wherein the reaction plate comprises a keyed registration feature. 
     
     
         17 . The system of  claim 1 , wherein the first manifold comprises a common interior lumen that is in fluid communication with all fluid in the manifold. 
     
     
         18 . The system of  claim 1 , wherein the reagent control system comprises a pressure selector. 
     
     
         19 . (canceled) 
     
     
         20 . The system of  claim 1 , wherein the system is configured to pneumatically drive all reagents from the reagent supply system to the subset of reaction vessels of the plurality of reaction vessels and then expel the reagents from the system through the outlet port. 
     
     
         21 . The system of  claim 1 , wherein, in operation, the plurality of reaction vessels are spatially fixed relative to the reagent supply system. 
     
     
         23 . (canceled) 
     
     
         24 . A computer-controlled system for performing a plurality of different reactions in parallel, each in a separate reaction vessel, wherein the system is configured to be in a closed configuration when one or more reagents are initially delivered to the reaction vessel. 
     
     
         25 . The system of  claim 24 , wherein the system is configured to prevent a first reagent of the one or more reagents from contact with moisture from the external environment prior to expulsion of the reagent from the system. 
     
     
         26 . The system of  claim 24 , wherein the one or more reagents comprises a plurality of reagents, and wherein the system is configured to prevent each of the plurality of reagents from contact with moisture from an external environment prior to expulsion of the reagent from the system. 
     
     
         27 . The system of  claim 24 , wherein the system is configured such that the water content of a first reagent of the one or more reagents never exceeds 10 ppm by weight until at least expulsion of the first reagent from the system. 
     
     
         28 . The system of  claim 24 , wherein the system is configured to pneumatically:
 (a) deliver the first reagent a subset of the reaction vessels;   (b) agitate the first reagent in each reaction vessel of the subset of the reaction vessels to promote mixing; and   (c) expel the first reagent from the subset of reaction vessels;   wherein (a), (b), and (c) occur without the first reagent contacting moisture from an external environment.   
     
     
         29 . The system of  claim 28 , wherein the system is configured to pneumatically deliver the first reagent to a subset of the reaction vessels without entering into reaction vessels that are not within the subset. 
     
     
         30 - 121 . (canceled) 
     
     
         122 . A method for synthesizing a plurality of different oligonucleotides that each comprise a first oligonucleotide segment and a second oligonucleotide segment, the method comprising synthesizing, in each of a plurality of reaction vessels, a second oligonucleotide segment onto a first oligonucleotide segment that is attached to solid support media, wherein first oligonucleotide segment is identical between each of the plurality of different oligonucleotides, and the second oligonucleotide segment differs between each of the plurality of different oligonucleotides. 
     
     
         123 . The method of  claim 122 , further comprising, prior to synthesizing the second oligonucleotide segment onto the first oligonucleotide segment, transferring different portions of a single source of solid-phase media to each of the plurality of reaction vessels, wherein, at the time of transfer, the solid phase media is attached to the first oligonucleotide segment. 
     
     
         124 . (canceled) 
     
     
         125 . The method of  claim 122 , wherein the first oligonucleotide segment is synthesized on a first oligonucleotide synthesizer and the second oligonucleotide segment is synthesized on a second oligonucleotide synthesizer. 
     
     
         126 . The method of  claim 122 , wherein the plurality of reaction vessels are formed from a single reaction plate. 
     
     
         127 - 132 . (canceled) 
     
     
         133 . The method of  claim 122 , wherein the second oligonucleotide segment is between 2 and 150 nucleotides in length. 
     
     
         134 . The method of  claim 122 , wherein the second oligonucleotide segment is between 17 and 22 nucleotides in length. 
     
     
         135 - 153 . (canceled)