Methods for high fidelity production of long nucleic acid molecules
First Claim
1. A method for synthesizing a nucleic acid molecule, the method comprising:
- a) providing a first immobilized nucleic acid comprising a first 5′
region and a first 3′
region;
b) providing a second immobilized nucleic acid comprising a second 5′
region and a second 3′
region, wherein said second 3′
region and said first 5′
region comprise identical nucleic acid sequences;
c) hybridizing said first immobilized nucleic acid with an oligonucleotide under conditions promoting hybridization of said oligonucleotide to said first 3′
region, extending the hybridized oligonucleotide, and producing a first extension product comprising a 3′
region that is complementary to said first 5′
region; and
d) hybridizing said second immobilized nucleic acid with said first extension product under conditions promoting hybridization of said 3′
region of said first extension product to said second 3′
region, extending the 3′
region of said first extension product, and producing a second extension product comprising a 3′
region that is complementary to said second 5′
region, wherein said second extension product comprises a sequence complementary to said first 3′ and
5′
regions and said second 3′ and
5′
regions and said second extension product comprises the synthesized nucleic acid molecule.
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Abstract
In a method for synthesizing a long nucleic acid molecule, a first immobilized nucleic acid has a first 5′ region and a first 3′ region and a second immobilized nucleic acid has a second 5′ region and a second 3′ region. The second 3′ region and the first 5′ region have identical nucleic acid sequences. An oligonucleotide is hybridized to the first 3′ region, extending the hybridized oligonucleotide and producing a first extension product having a 3′ region that is complementary to the first 5′ region. The 3′ region of the first extension product is hybridized to the second 3′ region, extending the 3′ region of the first extension product and producing a synthesized nucleic acid molecule having a 3′ region that is complementary to the second 5′ region, wherein the synthesized nucleic acid molecule has a sequence complementary to the first and second 3′ and 5′ regions.
51 Citations
6 Claims
-
1. A method for synthesizing a nucleic acid molecule, the method comprising:
-
a) providing a first immobilized nucleic acid comprising a first 5′
region and a first 3′
region;b) providing a second immobilized nucleic acid comprising a second 5′
region and a second 3′
region, wherein said second 3′
region and said first 5′
region comprise identical nucleic acid sequences;c) hybridizing said first immobilized nucleic acid with an oligonucleotide under conditions promoting hybridization of said oligonucleotide to said first 3′
region, extending the hybridized oligonucleotide, and producing a first extension product comprising a 3′
region that is complementary to said first 5′
region; andd) hybridizing said second immobilized nucleic acid with said first extension product under conditions promoting hybridization of said 3′
region of said first extension product to said second 3′
region, extending the 3′
region of said first extension product, and producing a second extension product comprising a 3′
region that is complementary to said second 5′
region, wherein said second extension product comprises a sequence complementary to said first 3′ and
5′
regions and said second 3′ and
5′
regions and said second extension product comprises the synthesized nucleic acid molecule. - View Dependent Claims (2, 3, 4, 5, 6)
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Specification