Programmable oligonucleotide synthesis
First Claim
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1. A method for isolating an individual nucleic acid molecule having a desired correct sequence from a mixture of nucleic acids, comprising:
- (a) monoclonizing a mixture of nucleic acids on a solid reaction support;
(b) parallel sequencing the monoclonized nucleic acids of step (a),(c) localizing a nucleic acid molecule with the desired correct sequence in the nucleic acids of step (b),(d) isolating the nucleic acid molecule from step (c), and(e) optionally further processing of the isolated nucleic acid molecule obtained in step (d).
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Abstract
The invention relates to methods and devices for preparing synthetic nucleic acids.
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Citations
18 Claims
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1. A method for isolating an individual nucleic acid molecule having a desired correct sequence from a mixture of nucleic acids, comprising:
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(a) monoclonizing a mixture of nucleic acids on a solid reaction support; (b) parallel sequencing the monoclonized nucleic acids of step (a), (c) localizing a nucleic acid molecule with the desired correct sequence in the nucleic acids of step (b), (d) isolating the nucleic acid molecule from step (c), and (e) optionally further processing of the isolated nucleic acid molecule obtained in step (d). - View Dependent Claims (14, 15, 16)
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2. A method for isolating an individual nucleic acid having a desired sequence from a mixture of nucleic acids, comprising:
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(a) monoclonizing a mixture of nucleic acids on a solid reaction support; (b) parallel sequencing the monoclonized nucleic acids of step (a), (c) localizing a nucleic acid with the desired sequence in the nucleic acids of step (b), (d) isolating the nucleic acid from step (c), and (e) optionally further processing of isolated nucleic acid obtained in step (d) wherein the isolation takes place by at least one of steps (i) to (iv); (i) isolating one or more beads on which said nucleic acid molecule is immobilized, (ii) cleaving off the nucleic acid molecule from a support on which said nucleic acid molecule is immobilized, (iii) selectively amplifying the nucleic acid molecule by spatially-resolved addition of PCR reagents, (iv) eluting the nucleic acid molecule by laser capture method.
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3. A method for isolating an individual nucleic acid molecule having a desired correct sequence from a mixture of nucleic acids, comprising:
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(a) monoclonizing a plurality of the different nucleic acids by emulsion-based PCR on beads (b) immobilizing the beads in a reaction support (c) parallel sequencing the monoclonized nucleic acids of step (a) (d) localizing an individual nucleic acid molecule with the desired correct sequence in the nucleic acids of step (c) (e) isolating the beads comprising the nucleic acid molecules localized in step (d) (f) obtaining the clonal nucleic acid sequences by detachment of the clonal nucleic acid sequence from the bead or by copying (f) optionally further processing of the isolated nucleic acid molecule obtained in step (d).
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4. A method for quality control of assembled nucleic acid molecules, said method comprising:
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(a) providing a mixture of assembled nucleic acid molecules suspected of containing sequence defects, (b) amplifying the mixture of (a) to receive a clonal population thereof, (c) immobilizing the clonal population of (b) on a support, (d) parallel sequencing the clonal population immobilized according to (c), (e) localizing at least one clonal nucleic acid molecule that meets a predefined criterion (f) obtaining the at least one clonal nucleic acid molecule of (e). - View Dependent Claims (6, 7, 8, 9, 10, 11, 12, 13)
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5. A method for isolation of a sequence-verified nucleic acid clone, comprising:
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(a) providing a mixture of nucleic acid molecules suspected of containing sequence defects, (b) amplifying the mixture of (a) to receive a clonal population thereof, (c) immobilizing the clonal population of (b) on a support, (d) parallel sequencing the clonal population immobilized according to (c) (e) localizing at least one clonal nucleic acid molecule that meets a predefined criterion, and (f) obtaining the at least one clonal nucleic acid molecule of (e). - View Dependent Claims (17, 18)
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Specification