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US4458066A
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Process for preparing polynucleotides
Assignee: University Patents Inc
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Filing: 1981-03-24
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Granted: 1984-07-03
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Expires: 2001-03-24
Abstract
Abstract New and useful intermediate nucleotides bound to an inorganic polymer support, including the preparation thereof, and processes for the conversion to oligonucleotides which are especially useful for the synthesis of polynucleotides, particularly ribonucleic (RNA) and deoxyribonucleic acids (DNA).
Inventors
- 👤 Marvin H. Caruthers
- 👤 Mark D. Matteucci
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-23 19:07
CPC Classifications
C; C07; C07H; C07H21/00; B; B01; B01J; B01J19/00; B01J19/0046; C07H19/00; C07H19/02; C07H19/04; B01J2219/00; B01J2219/00274; B01J2219/00277; B01J2219/00279; B01J2219/00281; B01J2219/00286; B01J2219/00351; B01J2219/00389; B01J2219/00497; B01J2219/005; B01J2219/00583; B01J2219/0059; B01J2219/00596; B01J2219/0068; B01J2219/00686; B01J2219/00718; B01J2219/0072; B01J2219/00722; C40; C40B; C40B40/00; C40B40/04; C40B40/06; C40B60/00; C40B60/14; Y; Y02; Y02P; Y02P20/00; Y02P20/50; Y02P20/582
Claims Analysis
Total Claims: 49
Claim 1
Independent Claim
64 words
A modified inorganic polymer represented by the formula: ##STR5## wherein P is an inorganic polymer linked to the 3' or 5' --O-- of the nucleoside through a base hydrolyzable covalent bond; R is H or a blocking group; R 1 is a hydrocarbyl radical containing up to 10 carbons; each B is a nucleoside or deoxynucleoside base; and each A is H or OR.
Claim 2
Dependent Claim
64 words
A modified inorganic polymer represented by the formula: ##STR5## wherein P is an inorganic polymer linked to the 3' or 5' --O-- of the nucleoside through a base hydrolyzable covalent bond; R is H or a blocking group; R 1 is a hydrocarbyl radical containing up to 10 carbons; each B is a nucleoside or deoxynucleoside base; and each A is H or OR.
Claim 3
Dependent Claim
13 words
The modified polymer according to claim 1 wherein R 1 is lower alkyl.
Claim 4
Dependent Claim
14 words
The modified polymer according to claim 1 wherein the polymer is a silica gel.
Claim 5
Dependent Claim
18 words
The modified polymer according to claim 3 wherein free silanol hydroxy groups on the silica gel are blocked.
Claim 6
Dependent Claim
17 words
The modified polymer according to claim 3 wherein the 5' --O-- blocking group is a trityl group.
Claim 7
Dependent Claim
26 words
The modified polymer according to claim 3 wherein the silica gel is macroporous and of a particle size ranging from about 5 to about 1,000 m.
Claim 8
Dependent Claim
57 words
A modified inorganic polymer represented by the formula: ##STR6## wherein P is an inorganic polymer linked to the 3' --O through an ester linkage; R is H or a blocking group; R 1 is a hydrocarbyl radical containing up to 10 carbons, each B is a nucleoside or deoxynucleoside base and each A is H or OR.
Claim 9
Dependent Claim
57 words
A modified inorganic polymer represented by the formula: ##STR6## wherein P is an inorganic polymer linked to the 3' --O through an ester linkage; R is H or a blocking group; R 1 is a hydrocarbyl radical containing up to 10 carbons, each B is a nucleoside or deoxynucleoside base and each A is H or OR.
Claim 10
Dependent Claim
18 words
The modified polymer according to claim 7 wherein the polymer is a silica gel and A is H.
Claim 11
Dependent Claim
18 words
The modified polymer according to claim 8 wherein free silanol hydroxy groups of the silica gel are blocked.
Claim 12
Dependent Claim
17 words
The modified polymer according to claim 8 wherein the 5' --O-- blocking group is a trityl group.
Claim 13
Dependent Claim
15 words
The modified polymer according to claim 10 wherein the trityl group is dimethoxytrityl or monomethoxytrityl.
Claim 14
Dependent Claim
20 words
The modified polymer according to claim 8 wherein B is at least one of adenine, guanine, cytosine, uracil and thymine.
Claim 15
Dependent Claim
56 words
A modified inorganic polymer comprising an oligonucleotide or deoxyoligonucleotide covalently bound to said polymer through an ester linkage formed through the 3' --OH of the initial nucleoside moiety, wherein the terminal nucleoside is attached through a phosphite or phosphate triester linkage to the oligonucleotide chain and the 5' --O-- substituent is H or a blocking group.
Claim 16
Dependent Claim
56 words
A modified inorganic polymer comprising an oligonucleotide or deoxyoligonucleotide covalently bound to said polymer through an ester linkage formed through the 3' --OH of the initial nucleoside moiety, wherein the terminal nucleoside is attached through a phosphite or phosphate triester linkage to the oligonucleotide chain and the 5' --O-- substituent is H or a blocking group.
Claim 17
Dependent Claim
14 words
The modified polymer according to claim 13 wherein the polymer is a silica gel.
Claim 18
Dependent Claim
19 words
The modified polymer according to claim 12 wherein the free silanol hydroxy groups on the silica gel are blocked.
Claim 19
Dependent Claim
17 words
The modified polymer according to claim 12 wherein the 5' --O-- blocking group is a trityl group.
Claim 20
Dependent Claim
42 words
The modified polymer according to claim 14 wherein the oligonucleotide or deoxyoligonucleotide is attached to said silica gel through the 3' --O-- linkage, --Si--Z--NHCO--Z-- 1 --CO-- wherein each of Z and Z 1 is a hydrocarbyl radical containing up to 10 carbons.
Claim 21
Dependent Claim
89 words
A process for production of oligonucleotides which comprises the step of condensing the 3' --OH or 5' --OH group of a nucleoside or oligonucleotide covalently linked to an inorganic polymer by a coupling agent through the 5' --O-- or 3' --O--, respectively, of said nucleoside or oligonucleotide with a phosphite compound of the formula: ##STR7## wherein R is a blocking group; B is a nucleoside or deoxynucleoside base; A is H or OR; R 1 is lower alkyl; and X is Cl or Br, or a secondary amino group.
Claim 22
Dependent Claim
89 words
A process for production of oligonucleotides which comprises the step of condensing the 3' --OH or 5' --OH group of a nucleoside or oligonucleotide covalently linked to an inorganic polymer by a coupling agent through the 5' --O-- or 3' --O--, respectively, of said nucleoside or oligonucleotide with a phosphite compound of the formula: ##STR7## wherein R is a blocking group; B is a nucleoside or deoxynucleoside base; A is H or OR; R 1 is lower alkyl; and X is Cl or Br, or a secondary amino group.
Claim 23
Dependent Claim
41 words
The process according to claim 18 wherein the phosphite compound is of the formula: ##STR8## wherein substituents R,R 1 , X, A and B are as previously defined and said condensation occurs through the 5' --OH of said nucleoside or oligonucleotide.
Claim 24
Dependent Claim
12 words
The process according to claim 19 wherein R 1 is lower alkyl.
Claim 25
Dependent Claim
14 words
The process according to claim 19 wherein the inorganic polymer is a silica gel.
Claim 26
Dependent Claim
17 words
The process according to claim 21 wherein A is H and R is a trityl blocking group.
Claim 27
Dependent Claim
23 words
The process according to claim 22 wherein the free silanol hydroxy groups on the silica gel are blocked prior to the condensation reaction.
Claim 28
Dependent Claim
12 words
The process according to claim 22 wherein the trityl group is dimethoxytrityl.
Claim 29
Dependent Claim
22 words
The process according to claim 21 wherein the silica gel is covalently linked to said nucleoside or oligonucleotide through an ester linkage.
Claim 30
Dependent Claim
20 words
The process according to claim 21 including the further step of oxidizing the resulting phosphite triester to a phosphate triester.
Claim 31
Dependent Claim
13 words
The process according to claim 26 wherein I 2 is the oxidizing agent.
Claim 32
Dependent Claim
35 words
The process according to claim 26 wherein the 5' --O-- blocking group of the product thus obtained is removed and the product is then condensed with a further nucleoside phosphite compound of the said formula.
Claim 33
Dependent Claim
45 words
The process according to claim 28 wherein (1) the product thus obtained is oxidized to obtain the corresponding phosphate triester; (2) the 5' --O-- blocking group is removed; and (3) the product is condensed with a further nucleoside or oligonucleoside phosphite of the said formula.
Claim 34
Dependent Claim
28 words
The process according to claim 29 wherein the 5' --O-- blocking group is a trityl group and the said group is removed by reaction with a Lewis acid.
Claim 35
Dependent Claim
23 words
The process according to claim 30 wherein the 5' --O-- blocking group is a dimethoxytrityl group and said Lewis acid is zinc bromide.
Claim 36
Dependent Claim
24 words
The process according to claim 29 wherein each of steps 1, 2 and 3 is repeated to successively add nucleosides to the oligonucleotide chain.
Claim 37
Dependent Claim
27 words
The process according to claim 32 wherein the oligonucleotide chain comprises from about 10 to about 30 nucleotides attached through the 5' --O and 3' --O positions.
Claim 38
Dependent Claim
98 words
A process for production of oligonucleotides which comprises the step of condensing a compound of the formula: ##STR9## wherein B is a nucleoside or deoxynucleoside base, A is H, OH or OR; and P is silica covalently bound to the nucleoside by the group --Z 1 NHCOZCO-- wherein Z and Z 1 are each alkylene of up to 5 carbon atoms, with a nucleoside phosphite compound of the formula: ##STR10## wherein A and B are as previously defined; R 1 is lower alkyl; R is a blocking group; and X is Cl or Br, or a secondary amino.
Claim 39
Dependent Claim
98 words
A process for production of oligonucleotides which comprises the step of condensing a compound of the formula: ##STR9## wherein B is a nucleoside or deoxynucleoside base, A is H, OH or OR; and P is silica covalently bound to the nucleoside by the group --Z 1 NHCOZCO-- wherein Z and Z 1 are each alkylene of up to 5 carbon atoms, with a nucleoside phosphite compound of the formula: ##STR10## wherein A and B are as previously defined; R 1 is lower alkyl; R is a blocking group; and X is Cl or Br, or a secondary amino.
Claim 40
Dependent Claim
19 words
The process according to claim 34 including the further step of oxidizing the phosphite triester to a phosphate triester.
Claim 41
Dependent Claim
13 words
The process according to claim 35 wherein I 2 is the oxidizing agent.
Claim 42
Dependent Claim
35 words
The process according to claim 35 wherein the 5' --O-- blocking group of the product thus obtained is removed and the product is then condensed with a further nucleoside phosphite compound of the said formula.
Claim 43
Dependent Claim
43 words
The process according to claim 37 wherein (1) the product thus obtained is oxidized to obtain the corresponding phosphate triester; (2) the 5' --O blocking group is removed; and (3) the product is condensed with a further nucleoside phosphite of the said formula.
Claim 44
Dependent Claim
24 words
The process according to claim 38 wherein each of steps 1, 2 and 3 are repeated to successively add nucleosides to the oligonucleotide chain.
Claim 45
Dependent Claim
27 words
The process according to claim 39 wherein the oligonucleotide chain comprises from about 10 to about 30 nucleosides attached through the 5' --O and 3' --O positions.
Claim 46
Dependent Claim
26 words
The process according to claim 39 wherein the phosphate triester product is hydrolyzed to form the oligonucleotide and silica gel, and the oligonucleotide is separated therefrom.
Claim 47
Dependent Claim
17 words
The process according to claim 41 including the further step of removing blocking groups from the oligonucleotide.
Claim 48
Dependent Claim
39 words
A process for producing polynucleotides which comprises the step of condensing the 3' --OH or 5' --OH of a nucleoside linked to a functionalized inorganic polymer with a compound which is the reaction product of nucleoside and blocked phosphodichloridite.
Claim 49
Dependent Claim
39 words
A process for producing polynucleotides which comprises the step of condensing the 3' --OH or 5' --OH of a nucleoside linked to a functionalized inorganic polymer with a compound which is the reaction product of nucleoside and blocked phosphodichloridite.
References Cited (Backward Citations)
No citation records indexed.
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1987-07-03 | 1988-01-02 | 1988-07-03 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1991-07-03 | 1992-01-02 | 1992-07-03 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1995-07-03 | 1996-01-02 | 1996-07-03 | $7700.00 | Window Closed / Lapsed |