Methods and platform for screening and selecting metabolites and their receptors

Abstract

Disclosed herein are methods and platforms using Receptor Compartmentalized Partnered Replication (CPR), in which the partner gene is a receptor, signal transduction pathway, or metabolic pathway that leads to the production of an effector molecule for the receptor or signal transduction pathway. The signal transduction pathway or receptor is coupled to the production of a thermostable polymerase. Emulsification of libraries of organisms with primers that can amplify the partner gene led to the selective amplification of those partner genes that were best able to produce the thermostable polymerase during thermal cycling of the emulsion.

1 - 27 . (canceled) 
     
     
         28 . A eukaryotic cell, wherein the cell comprises:
 a. an orthogonal signal transduction pathway or orthogonal receptor;   b. a polymerase coupled to said orthogonal signal transduction pathway or orthogonal receptor, wherein said polymerase is only produced when said orthogonal signal transduction pathway or orthogonal receptor is produced;   c. an effector molecule, wherein said effector molecule induces transcription of the orthogonal signal transduction pathway or orthogonal receptor, thereby also inducing expression of the polymerase.   
     
     
         29 . The eukaryotic cell of  claim 28 , wherein the cell is in an oil-in-emulsion bubble. 
     
     
         30 . The eukaryotic cell of  claim 28 , wherein the orthogonal signal transduction pathway or receptor has undergone multiple rounds of selection. 
     
     
         31 . The eukaryotic cell of  claim 28 , wherein said eukaryotic cell is a yeast cell. 
     
     
         32 . The eukaryotic cell of  claim 28 , wherein said polymerase is thermostable polymerase. 
     
     
         33 . The eukaryotic cell of  claim 28 , wherein said orthogonal signal transduction pathway or orthogonal receptor is non-naturally occurring in the eukaryotic cell. 
     
     
         34 . The eukaryotic cell of  claim 33 , wherein said orthogonal signal transduction pathway or orthogonal receptor is a variant of a naturally occurring receptor or signal transduction pathway of the eukaryotic cell. 
     
     
         35 . The eukaryotic cell of  claim 28 , wherein the effector activates the orthogonal signal transduction pathway or orthogonal receptor. 
     
     
         36 . The eukaryotic cell of  claim 28 , wherein the polymerase is a DNA or RNA polymerase. 
     
     
         37 . The eukaryotic cell of  claim 28 , wherein the effector molecule is an agonist or antagonist of the orthogonal signal transduction pathway or orthogonal receptor. 
     
     
         38 . The eukaryotic cell of  claim 28 , wherein the orthogonal signal transduction pathway or orthogonal receptor and polymerase are encoded by a plasmid. 
     
     
         39 . A platform comprising a recombinant molecule, wherein said recombinant molecule encodes an orthogonal signal transduction pathway or orthogonal receptor coupled to a polymerase, and a library of primers, wherein said primers are designed to amplify potential effector molecules of the signal transduction pathway or receptor. 
     
     
         40 . The platform of  claim 39 , wherein the polymerase is a DNA polymerase or RNA polymerase. 
     
     
         41 . The platform of  claim 39 , wherein the effector molecule is an agonist or antagonist. 
     
     
         42 . The platform of  claim 39 , wherein the recombinant molecule is a plasmid. 
     
     
         43 . The platform of  claim 39 , wherein the platform is within a eukaryotic cell. 
     
     
         44 . The platform of  claim 43 , wherein the cell is in an oil-in-emulsion bubble. 
     
     
         45 . A platform comprising a recombinant molecule, wherein said recombinant molecule encodes an orthogonal signal transduction pathway or orthogonal receptor coupled to a polymerase, and an effector molecule, is an agonist or antagonist of the orthogonal signal transduction pathway or orthogonal receptor. 
     
     
         46 . The platform of  claim 45 , wherein the polymerase is a DNA polymerase or RNA polymerase. 
     
     
         47 . The platform of  claim 45 , wherein the effector molecule is an agonist or antagonist.