Patent ID: 9217144
Filing Date: 2015-12-22
Classification: B01J,C12N,C12Q

Abstract:
1. A method for producing at least one polynucleotide having a predefined sequence, the method comprising the steps of: (a) providing a support having a plurality of discrete addressable features at predetermined locations of the support, the support comprising at least a first plurality of support-bound oligonucleotides at a first feature and a second plurality of support-bound oligonucleotides at a second feature, wherein the first and second pluralities of support-bound oligonucleotides are single-stranded oligonucleotides and have different predefined sequences, and the first plurality of support-bound oligonucleotides comprises a sequence region at its 5′ end that is the same as a sequence region of a 3′ end of the second plurality of support-bound oligonucleotides; (b) providing a plurality single-stranded oligonucleotides, wherein the 3′ end of the plurality of the single-stranded oligonucleotides is complementary to the 3′ end of the first plurality of support-bound oligonucleotides at selected addressable features; (c) hybridizing the plurality of single-stranded oligonucleotides to the first plurality of support-bound oligonucleotides at the first feature; (d) subjecting the plurality of single-stranded oligonucleotides to a chain extension reaction, thereby forming double-stranded extension products; (e) dissociating the double-stranded extension products, thereby producing a first plurality of single-stranded complementary oligonucleotides; (f) transferring, in a predetermined order, the first plurality of single-stranded complementary oligonucleotides in droplet volume from the first feature at a first predetermined location to the second feature at a second predetermined location of the support, thereby bringing into contact the first plurality of single-stranded complementary oligonucleotides with the second plurality of support-bound oligonucleotides; (g) annealing the first plurality of single-stranded complementary oligonucleotides to the second plurality of support-bound oligonucleotides at the second feature such that the second plurality of support-bound oligonucleotides serves as a template for extension of the first plurality of complementary oligonucleotides in a chain extension reaction, thereby producing the polynucleotide; and (h) optionally amplifying the polynucleotide.