Method for the synthesis of nucleotide of oligonucleotide phosphoramidites

Abstract

This invention presents novel methods for recovery of phosphoramidites from the waste products of oligonucleotide synthesis. The methods include reacting a tribromophenoxydichlorophosphorane with an H-phosphonate in the presence of an amine.

What is claimed is:  
     
         1 . A method for the preparation of a phosphoramidite comprising the steps of: 
 reacting a compound of formula:                          wherein:    Z is an intersugar linkage;    R 1  is a hydroxyl protecting group;    R 2  is H, OH, O-alkyl, O-alkylamino, O-alkylalkoxy, a polyether of formula (O-alkyl) m  where m is 1 to about 10, or a protected hydroxyl group;    R 3  is a phosphoryl protecting group;    B is a nucleobase; and    n is 0 to about 100;    with a compound of formula:                          wherein:    X 1  is Br or Cl;    X 2  and X 3  are, independently, H, Br or Cl;    x and y are each, independently, 2 or 3, and the sum of x and y is 5; and    contacting the product of the reaction with a compound of formula HN(Q) 2  to yield a phosphoramidite of formula:                          wherein each Q is independently alkyl having from 1 to 15 carbons, aryl having from 6 to 14 carbons or can form a heterocyclic ring having from 2 to 10 carbons with the nitrogen of the phosphoramidite.    
     
     
         2 . The method of  claim 1  wherein Q is isopropyl.  
     
     
         3 . The method of  claim 1  wherein R 2  is H.  
     
     
         4 . The method of  claim 1  wherein R 2  is a protected hydroxyl group.  
     
     
         5 . The method of  claim 1  wherein R 2  is O-alkyl or O-alkylalkoxy.  
     
     
         6 . The method of  claim 1  wherein R 2  is O-alkylalkoxy.  
     
     
         7 . The method of  claim 1  wherein R 2  is 2-methoxyethoxy.  
     
     
         8 . The method of  claim 1  wherein R 3 is cyanoethyl, 4-cyano-2-butenyl, or diphenylmethylsilylethyl.  
     
     
         9 . The method of  claim 1  wherein x is 2 and y is 3.  
     
     
         10 . The method of  claim 1  wherein x is 3 and y is 2.  
     
     
         11 . The method of  claim 1  wherein the reaction is performed in the presence of a solvent.  
     
     
         12 . The method of  claim 11  wherein the reaction is performed in the presence of a solvent is acetonitrile.  
     
     
         13 . The method of  claim 1  wherein the nucleobase is adenine, guanine, cytosine, thymine, uracil, 5-methyl cytosine or a protected derivative thereof.  
     
     
         14 . The method of  claim 1  wherein Z is a phosphodiester linkage, a phosphorothioate linkage, a phosphorodithioate linkage; or a phosphonate linkage.  
     
     
         15 . The method of  claim 14  wherein Z is a phosphodiester linkage or a phosphorothioate linkage.  
     
     
         16 . The method of  claim 1  wherein R 3  is cyanoethyl.  
     
     
         17 . The method of  claim 1  wherein R 3  is 4-cyano-2-butenyl.  
     
     
         18 . The method of  claim 1  wherein X 1 , X 2  and X 3  are Br located at 2,4 and 6-positions of the phenyl ring.