Methods and Assays for Capture of Nucleic Acids
First Claim
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1. A method for capturing target nucleic acid sequences comprising:
- a) providing;
i) a nucleic acid sample wherein said nucleic acid sample may or may not comprise a target sequence,ii) at least one flap endonuclease and at least one ligase, andiii) a plurality of oligonucleotide probes, wherein said probes comprise target sequences and a hairpin structure,b) applying said nucleic acid sample to said oligonucleotide probes under conditions wherein hybridization is allowed to occur between target sequences and probes; and
c) applying at least one flap endonuclease and at least one ligase to the hybridized nucleic acid/probes complex under conditions wherein enzymatic reactions are allowed to occur thereby capturing a nucleic acid target sequence.
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Abstract
The present disclosure provides methods and systems for sequence specific nucleic acid target capture comprising enzymatic reactions. The present disclosure relates to a plurality of oligonucleotide probes for capture and subsequent detection of target nucleic acid sequences, using flap endonucleases, ligases, and/or additional enzymes, proteins or compounds, on substrates, for example microarray slides, and in solution formats.
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Citations
28 Claims
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1. A method for capturing target nucleic acid sequences comprising:
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a) providing; i) a nucleic acid sample wherein said nucleic acid sample may or may not comprise a target sequence, ii) at least one flap endonuclease and at least one ligase, and iii) a plurality of oligonucleotide probes, wherein said probes comprise target sequences and a hairpin structure, b) applying said nucleic acid sample to said oligonucleotide probes under conditions wherein hybridization is allowed to occur between target sequences and probes; and c) applying at least one flap endonuclease and at least one ligase to the hybridized nucleic acid/probes complex under conditions wherein enzymatic reactions are allowed to occur thereby capturing a nucleic acid target sequence. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21)
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22. A method for capturing target nucleic acids comprising:
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a) providing; i) a nucleic acid sample wherein said nucleic acid sample comprises a detection moiety and may or may not comprise a target sequence and, ii) at least one flap endonuclease, at least one ligase, and iii) a plurality of oligonucleotide probes, wherein said probes comprise target sequences and a hairpin structure wherein said hairpin structure comprises at least one cleavable sequence, b) applying said nucleic acid sample to said oligonucleotide probes under conditions wherein hybridization is allowed to occur between target sequences and probes, c) applying at least one flap endonuclease and at least one ligase to the hybridized nucleic acid/probes complex under conditions wherein enzymatic reactions are allowed to occur thereby capturing a nucleic acid target sequence. - View Dependent Claims (23, 24, 25)
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26. A composition for sequence specific nucleic acid capture comprising a flap endonuclease, a ligase, and an oligonucleotide probe wherein said probe comprises a hairpin structure and a complementary target nucleic acid sequence.
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27. A kit for capturing and detecting nucleic acid sequences comprising:
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a) at least one flap endonuclease, b) at least one thermostable ligase, c) a plurality of oligonucleotide probes affixed to a substrate, and d) at least one buffer. - View Dependent Claims (28)
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Specification