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US 6,309,836 B1Granted
Compounds for protecting hydroxyls and methods for their use
OwnerMarek Kwiatkowskiinventor
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Abstract
A hydrocarbyldithiomethyl-modified compound of the Formula: R1—O—CH2—S—S—R2 or a salt thereof wherein R1 is an organic molecule and R2 is a hydrocarbyl is useful for protecting and/or blocking hydroxyl groups in organic molecules such as nucleotides. The hydrocarbyldithiomethyl-modified compounds can also be used for chemically synthesizing oligonucleotides and for sequencing nucleic acid compounds.
Claims14
A method for sequencing a nucleic acid comprising the steps of:
- a) contacting a target nucleic acid with a primer under conditions wherein said primer anneals to said target nucleic acid in a sequence specific manner and wherein at least a portion of said primer is complementary to a portion of said target nucleic acid;
- b) incorporating a hydrocarbyldithiomethyl-modified nucleotide into said primer;
- and
- c) detecting incorporation of said hydrocarbyldithiomethyl-modified nucleotide, wherein said hydrocarbyldithiomethyl-modified nucleotide is complementary to said target nucleic acid at said hydrocarbyldithiomethyl-modified nucleotide's site of incorporation thereby identifying the sequence of one nucleobase of said target nucleic acid.
1 more element
A method for sequencing a nucleic acid comprising the steps of:
- a) contacting a target nucleic acid with a primer under conditions wherein said primer anneals to said target nucleic acid in a sequence specific manner and wherein at least a portion of said primer is complementary to a portion of said target nucleic acid;
- b) incorporating a first 3′-hydrocarbyldithiomethyl-modified nucleotide into said primer;
- c) detecting said incorporation of said first 3′-hydrocarbyldithiomethyl-modified nucleotide thereby identifying the sequence of a nucleobase of said target nucleic acid;
- d) removing said hydrocarbyldithiomethyl group from said first incorporated hydrocarbyldithiomethyl-modified nucleotide to form a first elongated primer having a free hydroxyl group;
- e) incorporating a second 3′-hydrocarbyldithiomethyl-modified nucleotide into said first elongated primer;
- and
- f) detecting said second hydrocarbyldithiomethyl-modified nucleotide thereby identifying the sequence of another nucleobase of said target nucleic acid, wherein said first 3′-hydrocarbyldithiomethyl-modified nucleotide and said second 3′-hydrocarbyldithiomethyl-modified nucleotide are complementary to said target nucleic acid at each said nucleotide's site of incorporation.
4 more elements
A method for sequencing a nucleic acid comprising the steps of:
- a) providing a primer array comprising a plurality of sequencing primers;
- b) contacting a target nucleic acid with said primer array under conditions wherein said sequencing primers anneal to said target nucleic acid in a sequence specific manner thereby forming target-primer complexes between complementary portions of said sequencing primers and said target nucleic acid;
- c) incorporating a first 3′-hydrocarbyldithiomethyl-modified nucleotide into at least one sequencing primer portion of said target-primer complexes, said first 3′-hydrocarbyldithiomethyl-modified nucleotide being complementary to said target nucleic acid;
- and
- d) detecting said incorporation of said first 3′-hydrocarbyldithiomethyl-modified nucleotide, wherein said first 3′-hydrocarbyldithiomethyl-modified nucleotide is complementary to said target sequence at said first 3′-hydrocarbyldithiomethyl-modified nucleotide's site of incorporation.
2 more elements
Dependencies are inferred from the claim text.