System for rapid high-resolution gel electrophoresis

Abstract

Electrophoretic systems, formulations and methods are described which allow a user to perform electrophoresis experiments under conditions of high voltage and with reduced run time. An electrophoretic system, formulation or method may be run at 50% higher field strength than comparable systems already in use in the art. The presently described systems and formulations may be run at voltages above 225 V, above 250 V, above 275 V, above 300 V, above 325 V or above 350 V. The time required for performing an electrophoresis experiment may be reduced to less than about 30 minutes, less than about 20 minutes, less than about 15 minutes or less than about 12 minutes.

1 . A discontinuous buffer electrophoretic system comprising:
 an electrophoretic separation gel, said electrophoretic separation gel comprising a gel buffer system, said buffer system comprising:   a gel amine buffer; and   a gel ampholyte; and   an aqueous running buffer comprising a buffer ampholyte.   
     
     
         2 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the separation gel comprises acrylamide, agarose, or acrylamide and agarose. 
     
     
         3 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the separation gel comprises acrylamide. 
     
     
         4 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the separation gel comprises between about 6 to about 25 wt. % acrylamide. 
     
     
         5 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the separation gel comprises between about 8 to about 20 wt. % acrylamide. 
     
     
         6 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the separation gel comprises between about 8 to about 15 wt. % acrylamide. 
     
     
         7 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the electrophoretic separation gel further comprises a stacking gel coupled to one end thereof. 
     
     
         8 . The discontinuous buffer electrophoretic system according to  claim 7 , wherein the stacking gel comprises about 4 wt. % acrylamide. 
     
     
         9 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the gel ampholyte is different from the buffer ampholyte. 
     
     
         10 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pK of the gel amine buffer is in the range of about 5.5 to about 7.5. 
     
     
         11 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the gel amine buffer is Bis-Tris. 
     
     
         12 . The discontinuous buffer electrophoretic system according to either  claim 10  or  claim 11 , wherein the concentration of the gel amine buffer is in the range of about 190 mM to about 295 mM. 
     
     
         13 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the concentration of the gel amine buffer is less than about 300 mM. 
     
     
         14 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the concentration of the gel amine buffer is between about 175 mM to about 300 mM. 
     
     
         15 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pK of the gel ampholyte is about 1.5 pK units greater than the pK of the gel amine. 
     
     
         16 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pK of the gel ampholyte is between about 7 to about 9. 
     
     
         17 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pK of the gel ampholyte is between about 8 to about 8.5. 
     
     
         18 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pK of the gel ampholyte is between about 8.1 to about 8.3. 
     
     
         19 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the gel ampholyte is Tricine, Bicine, glycinamide or acetamidoglycine. 
     
     
         20 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the gel ampholyte is Tricine or Bicine. 
     
     
         21 . The discontinuous buffer electrophoretic system according to either of  claim 19  or  20 , wherein the concentration of the gel ampholyte is in the range of about 25 mM to about 125 mM. 
     
     
         22 . The discontinuous buffer electrophoretic system according to either of  claim 19  or  20 , wherein the concentration of the gel ampholyte is in the range of about 50 mM to about 100 mM. 
     
     
         23 . The discontinuous buffer electrophoretic system according to either of  claim 19  or  20 , wherein the concentration of the gel ampholyte is about 50 mM. 
     
     
         24 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pH of the gel buffer system is in the range of about 6 to about 8 
     
     
         25 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pH of the gel buffer system is in the range of about 6.0 to about 7.5. 
     
     
         26 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pH of the gel buffer system is about 6.5. 
     
     
         27 . The discontinuous buffer electrophoretic system according to any of  claims 24  to  26 , wherein the pH is adjusted by the addition of an acid. 
     
     
         28 . The discontinuous buffer electrophoretic system according to  claim 27 , wherein the acid is HCl. 
     
     
         29 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the gel buffer system further includes a source of Cl −  ions. 
     
     
         30 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the gel buffer system further includes about 95 mM to about 170 mM Cl −  ions. 
     
     
         31 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the gel buffer system further includes about 135 mM to about 145 mM Cl −  ions. 
     
     
         32 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the pK of the buffer ampholyte in the running buffer is less than the pK of the gel ampholyte. 
     
     
         33 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the buffer ampholyte comprises MES, ADA, PIPES, ACES, BES, TES, HEPES, MOPS, CAPSO, DIPSO, POPSO, HEPPS, HEPPSO, salts thereof, or any combination thereof. 
     
     
         34 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the buffer ampholyte comprises MES, MOPS or DIPSO. 
     
     
         35 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the buffer ampholyte comprises MES. 
     
     
         36 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the buffer ampholyte comprises MOPS. 
     
     
         37 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the buffer ampholyte comprises DIPSO. 
     
     
         38 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the electrophoretic separation gel further comprises sucrose. 
     
     
         39 . The discontinuous buffer electrophoretic system according to  claim 38 , wherein the concentration of sucrose is less than about 5 wt. %. 
     
     
         40 . The discontinuous buffer electrophoretic system according to  claim 38 , wherein the concentration of sucrose is less than about 3 wt. %. 
     
     
         41 . The discontinuous buffer electrophoretic system according to  claim 38 , wherein the concentration of sucrose is less than about 2 wt. %. 
     
     
         42 . The discontinuous buffer electrophoretic system according to  claim 38 , wherein the concentration of sucrose is less than about 1.5 wt. %. 
     
     
         43 . The discontinuous buffer electrophoretic system according to  claim 1 , further comprising SDS in at least the separating gel. 
     
     
         44 . The discontinuous buffer electrophoretic system according to  claim 1 , further comprising SDS in the separating gel and the running buffer. 
     
     
         45 . The discontinuous buffer electrophoretic system according to  claim 1 , wherein the separating gel is a slab gel. 
     
     
         46 . The discontinuous buffer electrophoretic system according to  claim 45 , wherein the slab gel is between two glass or plastic plates. 
     
     
         47 . A kit comprising, in at least a first suitable container:
 one or more gel cassettes, each of said gel cassettes comprising the discontinuous buffer electrophoretic system according to any of  claims 1  to  47 ; and   an aqueous running buffer according to any of  claims 1  to  47 .