Compositions and methods for analysis of nucleic acids
First Claim
Patent Images
1. A method of preparing a DNA molecule comprising:
- a) ligating an adaptor to the end of a template DNA molecule, wherein said ligation introduces a nick or gap site onto said template DNA molecule;
b) initiating a nick translation reaction at the nick or gap site of said adaptor using a DNA polymerase having 5′
to 3′
exonuclease activity;
c) allowing said nick translation reaction to proceed for a specific time to produce DNA fragments of similar size; and
d) terminating said nick translation reaction.
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Abstract
Disclosed are a number of methods that can be used in a variety of embodiments, including, creation of a nucleic acid terminated at one or more selected bases, sequence analysis of nucleic acids, mapping of sequence motifs within a nucleic acid, positional mapping of nucleic acid clones, and analysis of telomeric regions. The methods utilize double-stranded templates, and in most aspects involve a strand replacement reaction initiated at one or more random or specific locations created in a nucleic acid molecule, and in certain aspects utilizing an oligonucleotide primer.
168 Citations
46 Claims
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1. A method of preparing a DNA molecule comprising:
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a) ligating an adaptor to the end of a template DNA molecule, wherein said ligation introduces a nick or gap site onto said template DNA molecule;
b) initiating a nick translation reaction at the nick or gap site of said adaptor using a DNA polymerase having 5′
to 3′
exonuclease activity;
c) allowing said nick translation reaction to proceed for a specific time to produce DNA fragments of similar size; and
d) terminating said nick translation reaction. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24)
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25. A method of preparing a DNA molecule comprising:
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a) ligating an adaptor to the end of a template DNA molecule, wherein said ligation introduces a nick or gap site onto said template DNA molecule, said nick or gap site comprised in said adaptor;
b) initiating a nick translation reaction at the nick or gap site of said adaptor using a DNA polymerase having 5′
to 3′
exonuclease activity;
c) allowing said nick translation reaction to proceed for a specific time; and
d) terminating said nick translation reaction. - View Dependent Claims (26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46)
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Specification