Patent ID: 11060137

Abstract:
The invention provides compositions and methods for assembling a DNA molecule having a desired sequence. The methods involve contacting a DNA polymerase, dNTPs, and a plurality of pairs of oligonucleotides. The oligonucleotides of a pair have a portion of the desired sequence, and an internal sequence that overlaps and is complementary to an internal sequence of the other oligonucleotide of the pair, and, when arranged in order, they have at least a portion of the desired sequence. The oligonucleotides also have a 3′ or a 5′ primer binding sequence having a binding site for a primer. The oligonucleotides that correspond to the end oligonucleotides of the desired sequence also have a universal 3′ flanking sequence and a universal 5′ flanking sequence, respectively. The methods involve performing a first amplification reaction on the plurality of pairs of oligonucleotides; removing the 3′ and 5′ primer binding sequences from the plurality of pairs of oligonucleotides; and subjecting the plurality of pairs of oligonucleotides to an assembly reaction to thereby assemble the dsDNA molecule having the desired sequence.