Development of novel detergents for use in pcr systems

Abstract

This disclosure relates to novel detergents for use in various procedures including, for example, nucleic acid amplification reactions such as polymerase chain reaction (PCR). Methods for preparing the modified detergents are also described.

1 - 57 . (canceled) 
     
     
         58 . A compound of Formula I or an alkyl ester thereof: 
       
         
           
           
               
               
           
         
       
       wherein:
 R 1  is a (C 8 -C 16 ) alkyl or (C 8 -C 16 ) substituted alkyl; 
 R 2  and R 3  are each independently H or CH 3 ; 
 R 4  and R 5  are each independently H, (CH 2 ) n NH, (CH 2 ) n N, or, alternatively, R 3  is taken together with R 5  to form a 5- or 6-membered ring which is optionally substituted with at least one (C 1 -C 30 ) alkyl, (C 1 -C 30 ) substituted alkyl, (C 1 -C 30 ) heteroalkyl, or (C 1 -C 30 ) substituted heteroalkyl; and 
 each n is independently 1 to 30, 
 wherein when R 2 , R 3 , R 4 , and R 5  are H, R 1  is a C 8  alkyl or (C 8 -C 16 ) substituted alkyl, 
 wherein when R 2  and R 3  are CH 3  and R 4  and R 5  are H, R 1  is a C 8  alkyl or (C 8 -C 16 ) substituted alkyl, and 
 wherein when R 3  is taken together with R 5  to form a 5-membered ring and R 4  is H, R 1  is a (C 8 -C 16 ) substituted alkyl, R 2  is CH 3 . 
 
     
     
         59 . A nucleic acid amplification reaction mixture comprising:
 a) at least one polymerase;   b) dNTPs; and   c) at least one compound or alkyl ester thereof according to  claim 58 .   
     
     
         60 . The reaction mixture according to  claim 59 , wherein said polymerase is thermostable. 
     
     
         61 . The reaction mixture of  claim 59 , further comprising at least one nucleic acid primer. 
     
     
         62 . The reaction mixture of  claim 59 , further comprising a detectable label. 
     
     
         63 . The reaction mixture of  claim 62 , wherein the detectable label is part of a primer. 
     
     
         64 . The reaction mixture of  claim 62 , wherein the detectable label is part of a probe. 
     
     
         65 . The reaction mixture of  claim 60 , wherein the thermostable polymerase is selected from the group consisting of Taq DNA polymerase, Tfi DNA polymerase, Tfl DNA polymerase, Pfu DNA polymerase, and Vent™ DNA polymerase, a polymerase having reduced 3′-to-5′ exonuclease activity, SuperScript™ DNA polymerase, a genetically engineered DNA polymerase, a polymerase having the active site mutation F667Y, a polymerase having the equivalent of active site F667Y, Tth polymerase, AmpliTaq®FS, ThermoSequenase™, Therminator I, Therminator II, Therminator III, Therminator Gamma, a derivative thereof, and a fragment thereof. 
     
     
         66 . The reaction mixture of  claim 65 , wherein the thermostable polymerase is Taq DNA polymerase. 
     
     
         67 . The compound or alkyl ester thereof according to  claim 58 , wherein the alkyl ester is an ethyl ester. 
     
     
         68 . A compound of Formula I: 
       
         
           
           
               
               
           
         
       
       wherein:
 R 1  is a (C 8 -C 16 ) alkyl or (C 8 -C 16 ) substituted alkyl; 
 R 2  and R 3  are each independently H or CH 3 ; 
 R 4  and R 5  are each independently H, (CH 2 ) n NH, (CH 2 ) n N, or, alternatively, R 3  is taken together with R 5  to form a 5- or 6-membered ring which is optionally substituted with at least one (C 1 -C 30 ) alkyl, (C 1 -C 30 ) substituted alkyl, (C 1 -C 30 ) heteroalkyl, or (C 1 -C 30 ) substituted heteroalkyl; and 
 each n is independently 1 to 30, 
 with the proviso that the compound is not one of 
 
       
         
           
           
               
               
           
         
       
     
     
         69 . A nucleic acid amplification reaction mixture comprising:
 a) at least one polymerase;   b) dNTPs; and   c) at least one compound or alkyl ester thereof according to  claim 68 .   
     
     
         70 . The reaction mixture according to  claim 69 , wherein said polymerase is thermostable. 
     
     
         71 . The reaction mixture of  claim 69 , further comprising at least one nucleic acid primer. 
     
     
         72 . The reaction mixture of  claim 69 , further comprising a detectable label. 
     
     
         73 . The reaction mixture of  claim 72 , wherein the detectable label is part of a primer. 
     
     
         74 . The reaction mixture of  claim 72 , wherein the detectable label is part of a probe. 
     
     
         75 . The reaction mixture of  claim 70 , wherein the thermostable polymerase is selected from the group consisting of Taq DNA polymerase, Tfi DNA polymerase, Tfl DNA polymerase, Pfu DNA polymerase, and Vent™ DNA polymerase, a polymerase having reduced 3′-to-5′ exonuclease activity, SuperScript™ DNA polymerase, a genetically engineered DNA polymerase, a polymerase having the active site mutation F667Y, a polymerase having the equivalent of active site F667Y, Tth polymerase, AmpliTaq®FS, ThermoSequenase™, Therminator I, Therminator II, Therminator III, Therminator Gamma, a derivative thereof, and a fragment thereof. 
     
     
         76 . The reaction mixture of  claim 75 , wherein the thermostable polymerase is Taq DNA polymerase. 
     
     
         77 . The compound or alkyl ester thereof according to  claim 68 , wherein the alkyl ester is an ethyl ester.