Patent ID: 8808986
Filing Date: 2014-08-19
Classification: B01J,B01L,C12N,C12Q

Abstract:
1. A method for assembling a polynucleotide having a predefined sequence from a plurality of different oligonucleotides, the method comprising: a) providing a plurality of single-stranded template oligonucleotides at different features on a solid support, wherein each of the plurality of template oligonucleotides comprises a predefined sequence, a primer binding site and a sequence region that is the same or complementary to a sequence region of a single-stranded oligonucleotide present at a different feature of the solid support; b) generating a complementary oligonucleotide for each of the plurality of template oligonucleotides by enzyme-catalyzed synthesis within a primary droplet, thereby producing a plurality of double-stranded oligonucleotides; c) releasing the complementary oligonucleotides from the double-stranded oligonucleotides into the primary droplet; d) combining at least a first and second primary droplets on the support, thereby forming a secondary droplet, wherein the first primary droplet includes a released oligonucleotide that comprises a sequence region that is complementary to a sequence region of a released or template oligonucleotide from the second primary droplet; and e) exposing the secondary droplet to conditions suitable for hybridization and ligation, polymerase extension, or polymerase extension and ligation to assemble a double-stranded polynucleotide having a predefined sequence.