Base-modified nucleotides and cleavage of polynucleotides incorporating them
First Claim
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1. A method for cleaving a polynucleotide, comprising:
- replacing a natural nucleotide at greater than 90% of its points of occurrence in a polynucleotide with a base-modified nucleotide to form a base-modified polynucleotide; and
, contacting the base-modified polynucleotide with a reagent or combination of reagents that cleaves it at greater than 90% of the points of occurrence of the base-modified nucleotide, wherein;
the base-modified nucleotide has the chemical structure;
wherein R is a ribose or 2′
-deoxyribose moiety of an oligonucleotide or of a polynucleotide;
R1 and R2 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, aralkyl and alkaryl, wherein, if R1 or R2 contains two or more contiguous methylene (—
CH2—
) groups, any two such methylene groups may have interjected between them another group selected from the group consisting of —
O—
, —
C(O)NH—
, —
C(O)NHC(O)—
, —
NH—
, —
C(S)NH—
, —
CO—
, —
CS—
, —
S— and
(—
CF2—
)m, wherein m is 1-10;
R3 is hydrogen or —
NH2; and
, n is 0, 1 or 2.
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Abstract
The present invention is directed to novel base-modified nucleotides and methods for their use in the preparation and cleavage of modified polynucleotides.
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Citations
15 Claims
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1. A method for cleaving a polynucleotide, comprising:
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replacing a natural nucleotide at greater than 90% of its points of occurrence in a polynucleotide with a base-modified nucleotide to form a base-modified polynucleotide; and
,contacting the base-modified polynucleotide with a reagent or combination of reagents that cleaves it at greater than 90% of the points of occurrence of the base-modified nucleotide, wherein;
the base-modified nucleotide has the chemical structure;
whereinR is a ribose or 2′
-deoxyribose moiety of an oligonucleotide or of a polynucleotide;
R1 and R2 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, aralkyl and alkaryl, wherein, if R1 or R2 contains two or more contiguous methylene (—
CH2—
) groups, any two such methylene groups may have interjected between them another group selected from the group consisting of —
O—
, —
C(O)NH—
, —
C(O)NHC(O)—
, —
NH—
, —
C(S)NH—
, —
CO—
, —
CS—
, —
S— and
(—
CF2—
)m, wherein m is 1-10;
R3 is hydrogen or —
NH2; and
,n is 0, 1 or 2. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 14, 15)
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9. A method for cleaving a polynucleotide, comprising:
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replacing a natural nucleotide at greater than 90% of its points of occurrence in a polynucleotide with a base-modified nucleotide to form a base-modified polynucleotide; and
,contacting the base modified polynucleotide with a secondary amine having a boiling point greater than 100°
C. at atmospheric pressure whereby the polynucleotide is cleaved at greater than 90% of the sites of incorporation of the modified nucleotide.- View Dependent Claims (10, 11, 12, 13)
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Specification