Abstract: Methods for making a synthetic nucleic acid which comprise: (a) identifying a conflicting nucleotide sequence in a target sequence; (b) inserting a masking sequence into the conflicting sequence to produce a disrupted target sequence, wherein: (i) the masking sequence comprises recognition sites for one or more Type IIS restriction endonucleases; and (ii) digestion of said disrupted target sequence by said one or more Type IIS restriction endonucleases followed by re-ligation reconstitutes the target sequence; (c) synthesizing a polynucleotide comprising the disrupted target sequence using polymerase chain assembly; and (d) removing the masking sequence from said polynucleotide by digesting said polynucleotide with said one or more Type IIS restriction endonucleases followed by re-ligation of the digestion product, thereby producing a polynucleotide comprising said target sequence.

Abstract: This disclosure provides a method comprising: a) clamping the top and bottom strands of a double stranded DNA molecule to produce a duplex in which the top and bottom strands are linked; b) denaturing the duplex to produce a denatured product; and c) renaturing the denatured product in the presence of a labeled oligonucleotide that is complementary to a sequence of nucleotides in the double stranded DNA molecule, thereby producing a D-loop-containing product. Kits for performing the method and products made by the method are also provided.

Abstract: Methods for making a synthetic nucleic acid which comprise: (a) identifying a conflicting nucleotide sequence in a target sequence; (b) inserting a masking sequence into the conflicting sequence to produce a disrupted target sequence, wherein: (i) the masking sequence comprises recognition sites for one or more Type IIS restriction endonucleases; and (ii) digestion of said disrupted target sequence by said one or more Type IIS restriction endonucleases followed by re-ligation reconstitutes the target sequence; (c) synthesizing a polynucleotide comprising the disrupted target sequence using polymerase chain assembly; and (d) removing the masking sequence from said polynucleotide by digesting said polynucleotide with said one or more Type IIS restriction endonucleases followed by re-ligation of the digestion product, thereby producing a polynucleotide comprising said target sequence.

Abstract: Provided herein is a method comprising: producing a first complex by annealing: a first nucleic acid comprising, in order, a first unique sequence, a first central sequence and a second unique sequence; and a second nucleic acid comprising, in order, said second unique sequence, a second central sequence and a third unique sequence; wherein the first, second and third unique sequences do not hybridize with each other; subjecting said first complex to multiple rounds of primer extension to extend the first and second nucleic acids using each other as a template, thereby producing a first product molecule that contains, in order, the first unique sequence, the first central sequence, the second unique sequence, the second central sequence and the third unique sequence; and circularizing said first product molecule by intramolecularly ligating the ends of said product molecule together. Kits and compositions relating to the method are also provided.

Abstract: Provided herein is a method comprising: producing a first complex by annealing: a first nucleic acid comprising, in order, a first unique sequence, a first central sequence and a second unique sequence; and a second nucleic acid comprising, in order, said second unique sequence, a second central sequence and a third unique sequence; wherein the first, second and third unique sequences do not hybridize with each other; subjecting said first complex to multiple rounds of primer extension to extend the first and second nucleic acids using each other as a template, thereby producing a first product molecule that contains, in order, the first unique sequence, the first central sequence, the second unique sequence, the second central sequence and the third unique sequence; and circularizing said first product molecule by intramolecularly ligating the ends of said product molecule together. Kits and compositions relating to the method are also provided.

Abstract: This disclosure provides a method comprising: a) clamping the top and bottom strands of a double stranded DNA molecule to produce a duplex in which the top and bottom strands are linked; b) denaturing the duplex to produce a denatured product; and c) renaturing the denatured product in the presence of a labeled oligonucleotide that is complementary to a sequence of nucleotides in the double stranded DNA molecule, thereby producing a D-loop-containing product. Kits for performing the method and products made by the method are also provided.