Protein activity screening of clones having DNA from uncultivated microorganisms

Abstract

Disclosed is a process of screening clones having DNA from an uncultivated microorganism for a specified protein, e.g. enzyme, activity by screening for a specified protein, e.g. enzyme, activity in a library of clones prepared by (i) recovering DNA from a DNA population derived from at least one uncultivated microorganism; and (ii) transforming a host with recovered DNA to produce a library of clones which is screened for the specified protein, e.g. enzyme, activity.

What is claimed is:  
     
         1 . A method for identifying an enzymatic activity of interest comprising: 
 culturing a gene expression library comprising a pool of expression constructs, each expression construct comprising a vector containing one or more cDNA or genomic DNA fragments, wherein the cDNA or genomic DNA fragments in the pool of expression constructs are derived from a plurality of species of donor eukaryotic organisms; and    detecting the enzymatic activity encoded by the cDNA or genomic DNA fragments.    
     
     
         2 . The method of  claim 1 , wherein the enzymatic activity is selected from the group consisting of oxidoreductase, transferase, hydrolase, lyase, isomerase, and ligase activity.  
     
     
         3 . The method of  claim 1 , wherein the donor eukaryotic organisms are microorganisms.  
     
     
         4 . The method of  claim 3 , wherein the microorganisms are derived from an environmental sample.  
     
     
         5 . The method of  claim 1 , wherein the microorganisms are a mixed population of uncultured organisms.  
     
     
         6 . The method of  claim 1 , wherein the organisms are fungi.  
     
     
         7 . The method of  claim 1 , wherein the organisms are algae.  
     
     
         8 . The method of  claim 1 , wherein the organisms are protozoan.  
     
     
         9 . The method of  claim 5 , wherein the organisms are extremophiles.  
     
     
         10 . The method of  claim 9 , wherein the organisms are thermophiles, hyperthermophiles, psychrophiles, or psychrotrophs.  
     
     
         11 . The method of  claim 1 , wherein the host cell is a bacterial cell.  
     
     
         12 . The method of  claim 11 , wherein the bacterial cell is an  E. coli , Bacillus, Streptomyces, or  Salmonella typhimurium  cell.  
     
     
         13 . The method of  claim 1 , wherein the host cell is a fungal cell.  
     
     
         14 . The method of  claim 13 , wherein the fungal cell is a yeast cell.  
     
     
         15 . The method of  claim 1 , wherein the host cell is a Drosophila S2 or a Spodoptera S9 cell.  
     
     
         16 . The method of  claim 1 , wherein the host cell is an animal cell.  
     
     
         17 . The method of  claim 16 , wherein the animal cell is a CHO, COS or Bowes melanoma cell.  
     
     
         18 . The method of  claim 1 , wherein the host organism is a plant cell.  
     
     
         19 . A method for identifying an enzymatic activity of interest comprising: 
 culturing a gene expression library comprising a pool of expression constructs, each expression construct comprising a vector containing one or more cDNA or genomic DNA fragments, wherein the cDNA or genomic DNA fragments in the pool of expression constructs are derived from a plurality of species of donor organisms wherein the host cell is a bacterial cell; and    detecting the enzymatic activity encoded by the cDNA or genomic DNA fragments.    
     
     
         20 . A method for identifying an enzymatic activity of interest comprising: 
 culturing a gene expression library comprising a pool of expression constructs, each expression construct comprising a vector containing one or more cDNA or genomic DNA fragments, wherein the cDNA or genomic DNA fragments in the pool of expression constructs are derived from a plurality of species of donor organisms, wherein the host cell is a fungal cell; and    detecting the enzymatic activity encoded by the cDNA or genomic DNA fragments.    
     
     
         21 . A method for identifying an enzymatic activity of interest comprising: 
 culturing a gene expression library comprising a pool of expression constructs, each expression construct comprising a vector containing one or more cDNA or genomic DNA fragments, wherein the cDNA or genomic DNA fragments in the pool of expression constructs are derived from a plurality of species of donor organisms, wherein the host cell is a plant cell; and    detecting the enzymatic activity encoded by the cDNA or genomic DNA fragments.    
     
     
         22 . A method for identifying an enzymatic activity of interest comprising: 
 culturing a gene expression library comprising a pool of expression constructs, each expression construct comprising a vector containing one or more cDNA or genomic DNA fragments, wherein the cDNA or genomic DNA fragments in the pool of expression constructs are derived from a plurality of species of donor organisms wherein the host cell is an animal cell; and    detecting the enzymatic activity encoded by the cDNA or genomic DNA fragments.