Compositions and methods for selection of nucleic acids
First Claim
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1. A method for enrichment of a target region in a nucleic acid sample comprising:
- a) digesting the nucleic acid sample with a restriction enzyme that cuts a defined distance from its recognition site to produce a population of double-stranded nucleic acid fragments, wherein fragments containing the target region have known single-stranded overhangs on each end, each overhang being different;
b) ligating two types of stem-loop adapters to the population of nucleic acid fragments, wherein one type of stem-loop adapter has a single-stranded overhang sequence complementary to a first of the known single-strand overhangs at one end of the fragment comprising the target region, and the other type of stem-loop adapter has a single-stranded overhang sequence complementary to a second of the known single-stranded overhangs on the other end of the fragment comprising the target region;
c) treating the sample with one or more exonucleases to digest the double-stranded nucleic acid fragments that have one or no stem-loop adapter linked thereto, thus enriching for the target region in the nucleic acid sample.
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Abstract
Methods are provided for reducing the complexity of a population of nucleic acids prior to performing an analysis of the nucleic acids, e.g., sequence analysis. The methods result in a subset of the initial population enriched for a target region, which is typically located within one or more target fragments. The methods are particularly useful for analyzing populations having a high degree of complexity, e.g., chromosomal-derived DNA, whole genomic DNA, or mRNA populations.
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Citations
13 Claims
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1. A method for enrichment of a target region in a nucleic acid sample comprising:
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a) digesting the nucleic acid sample with a restriction enzyme that cuts a defined distance from its recognition site to produce a population of double-stranded nucleic acid fragments, wherein fragments containing the target region have known single-stranded overhangs on each end, each overhang being different; b) ligating two types of stem-loop adapters to the population of nucleic acid fragments, wherein one type of stem-loop adapter has a single-stranded overhang sequence complementary to a first of the known single-strand overhangs at one end of the fragment comprising the target region, and the other type of stem-loop adapter has a single-stranded overhang sequence complementary to a second of the known single-stranded overhangs on the other end of the fragment comprising the target region; c) treating the sample with one or more exonucleases to digest the double-stranded nucleic acid fragments that have one or no stem-loop adapter linked thereto, thus enriching for the target region in the nucleic acid sample. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13)
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Specification