Compositions and methods for the use of FMOC derivatives in DNA/RNA synthesis

• US 20040180355A1
• Filed: 09/24/2003
• Published: 09/16/2004
• Est. Priority Date: 09/27/2002
• Status: Active Grant

• Current Assignee: Gabriel Alvarado
• Original Assignee: Syngen Incorporated
• Inventors: Alvarado, Gabriel
• Granted Patent: US 7,534,872 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07H 21/02 with ribosyl as saccharide radical
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 2525/117 incorporating modified base

Abstract

Methods of nucleic acid preparation are described, including preparation of MnRNA, using FMOC derivatives to synthesize oligonucleotides in addition to applying FMOC protocols to various therapeutic and diagnostic methods. In some embodiments a single stranded oligonucleotide is synthesized bound to a polymer support (such as optic fiber glass filters) using said FMOC derivatives.

Claims

1. A method for synthesizing and purifying an oligonucleotide immobilized on a solid support complex, comprising:

a) providing;

i) a polymer support, and

ii) four pools of deoxyribonucleoside 3′

-phosphoramidite monomers in solution, wherein at least one of said monomers contains 9-fluorenylmethoxycarbonyl as an amino protecting group;

b) sequentially contacting said polymer support with a solution of monomers from one or more of said four pools under conditions such that a sequence of monomers is immobilized on said polymer support to generate an oligonucleotide/polymer support complex, wherein said complex comprises at least one monomer containing 9-fluorenylmethoxycarbonyl and wherein said solution, after said contacting, comprises unreacted material;

c) treating said complex under conditions such that said unreacted material is substantially removed, thereby creating a purified oligonucleotide/polymer support complex, and

d) treating said purified oligonucleotide/polymer support complex under conditions whereby said 9-fluorenylmethoxycarbonyl is released.

2. The method of claim 1, wherein said deoxyribonucleoside 3′

-phosphoramidite monomer containing 9-fluorenylmethoxycarbonyl as an amino protecting group is a 9-fluorenylmethoxycarbonyl-2′

-deoxycytidine 3′

-phosphoramidite.

3. The method of claim 1, wherein said deoxyribonucleoside 3′

-phosphoramidite monomer containing 9-fluorenylmethoxycarbonyl as an amino protecting group is a 9-fluorenylmethoxycarbonyl-2′

-deoxyadenosine 3′

-phosphoramidite.

4. The method of claim 1, wherein said deoxyribonucleoside 3′

-phosphoramidite monomer containing 9-fluorenylmethoxycarbonyl as an amino protecting group is a 9-fluorenylmethoxycarbonyl-2′

-deoxyguanosine 3′

-phosphoramidite.

5. The methods of claim 1, wherein the number of said monomers immobilized on said polymer support are in a range between one hundred and three hundred monomers.