In vitro evolution in microfluidic systems

Abstract

The invention describes a method for isolating one or more genetic elements encoding a gene product having a desired activity, comprising the steps of: (a) compartmentalising genetic elements into microcapsules; and (b) sorting the genetic elements which express the gene product having the desired activity; wherein at least one step is under microfluidic control. The invention enables the in vitro evolution of nucleic acids and proteins by repeated mutagenesis and iterative applications of the method of the invention.

1 - 107 . (canceled) 
     
     
         108 . A method for preparing an entity for analysis, the method comprising:
 obtaining a first fluid comprising a plurality of entities; and   forming droplets of the first fluid, each comprising multiple copies of a subset of said plurality, in a channel that contains droplets of a second fluid flowing in a third fluid, wherein the first, second, and third fluids are immiscible with each other.   
     
     
         109 . The method according to  claim 108 , wherein the first fluid is an aqueous fluid and the second and third fluids are different oils. 
     
     
         110 . The method according to  claim 109 , wherein the third fluid is a fluorinated or perfluorinated oil. 
     
     
         111 . The method according to  claim 108 , wherein the droplets of the first fluid are separated by droplets of the second fluid. 
     
     
         112 . The method according to  claim 108 , wherein the entities are nucleic acids, proteins, or cells. 
     
     
         113 . The method according to  claim 108 , wherein the entities are labeled. 
     
     
         114 . The method of  claim 113 , wherein the entities are optically labeled. 
     
     
         115 . The method of  claim 114 , wherein the entities are fluorescently labeled antibodies. 
     
     
         116 . The method of  claim 113 , wherein the entities are chemically labeled antibodies. 
     
     
         117 . The method of  claim 108 , wherein the first and second droplets are monodisperse with respect to each other. 
     
     
         118 . The method according to  claim 108 , wherein the entities are attached to microbeads. 
     
     
         119 . The method according to  claim 108 , wherein at least one of the first droplets includes only a single type of entity, present in multiple copies. 
     
     
         120 . The method according to  claim 108 , further comprising conducting a reaction in the first droplets. 
     
     
         121 . The method according to  claim 120 , further comprising detecting a reaction product in the first droplets. 
     
     
         122 . The method according to  claim 121 , wherein said detecting step comprises optically detecting.