Method of preparing libraries of template polynucleotides
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Abstract
The present invention relates to a method for preparing a library of template polynucleotides and use thereof in methods of solid-phase nucleic acid amplification. More specifically, the invention relates to a method for preparing a library of template polynucleotides that have common sequences at their 5′ ends and at their 3′ ends.
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Citations
58 Claims
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1-29. -29. (canceled)
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30. A method of generating a library of different polynucleotide molecules, the method comprising:
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(a) providing a plurality of different target polynucleotide duplexes; (b) providing identical polynucleotide adapters, wherein each adapter comprises a double-stranded annealed region and a mismatched region; (c) ligating the double-stranded annealed regions of the identical polynucleotide adapters to both ends of the different target polynucleotide duplexes to form adapter-target constructs, and (d) annealing a single universal primer species to the mismatched regions of the adapter-target constructs and extending the primer to form extension products complementary to both strands of the adapter-target constructs, wherein the extension products comprise nucleic acid sequences that differ from the nucleic acid sequences of either strand of the adapter-target constructs and the extension products collectively provide a library of different template polynucleotide molecules which have common sequences at their 5′
ends and common sequences at their 3′
ends,wherein the common sequences at the 5′
ends of the extension products are not complementary to the common sequences at the 3′
ends of the extension products. - View Dependent Claims (31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58)
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Specification