Induction of Gene Expression Using a High Concentration Sugar Mixture

Abstract

Described herein is a composition useful for inducing expression of genes whose expression is under control of an inducible promoter sequence and methods for the compositions preparation and use.

1 . A method of producing an inducing feed composition, said method comprising the steps of:
 a. Mixing a first solution with a whole cellulase preparation to give a first mixture; and   b. Incubating the first mixture at a temperature and for a sufficient time to produce the inducing feed composition.   
   
   
       2 . The method of  claim 1  wherein the first solution is a concentrated glucose solution comprising from about 5% to about 75% (wt/wt) glucose. 
   
   
       3 . The method of  claim 1  wherein the first solution is a concentrated glucose solution comprising from about 50% to about 75% (wt/wt) glucose. 
   
   
       4 . The method of  claim 1  wherein the first solution is a cellobiose solution comprising from about 5% to about 40% (wt/wt) cellobiose. 
   
   
       5 . The method of  claim 1  wherein the first solution is a cellobiose solution comprising from about 20% to about 40% (wt/wt) cellobiose. 
   
   
       6 . The method of  claim 1  wherein the whole cellulase preparation is from about 2 g/L to about 10 g/L protein. 
   
   
       7 . The method of  claim 1  wherein the whole cellulase preparation is about 5 g/L protein. 
   
   
       8 . The method of  claim 1  wherein the temperature is from about 50° C. to about 75° C. 
   
   
       9 . The method of  claim 1  wherein the solution is incubated for between 8 hours and 500 hours. 
   
   
       10 . The method of  claim 1  wherein the solution is incubated for between 48 hours and 72 hours. 
   
   
       11 . An inducing feed composition produced by the method of  claim 1 . 
   
   
       12 . The inducing feed composition of  claim 11  comprising a mixture of sugars. 
   
   
       13 . The inducing feed composition of  claim 11  comprising sophorose. 
   
   
       14 . The inducing feed composition of  claim 11  comprising gentiobiose. 
   
   
       15 . A method for producing proteins comprising providing a host cell with the inducing feed composition of  claim 11 . 
   
   
       16 . The method of  claim 15  wherein the protein produced is an endogenous cellulase. 
   
   
       17 . The method of  claim 15  wherein the host cell has been transformed with an expression construct comprising a promoter operably linked to a gene encoding a protein of interest. 
   
   
       18 . The method of  claim 17  wherein the promoter is an inducible promoter. 
   
   
       19 . The method of  claim 17  wherein the promoter is a cellulase gene promoter. 
   
   
       20 . The method of  claim 19  wherein the promoter is the cbh 1 promoter from  Trichoderma reesei.    
   
   
       21 . The method of  claim 18  wherein the inducible promoter is a sophorose-inducible promoter. 
   
   
       22 . The method of  claim 18  wherein the inducible promoter is a gentiobiose-inducible promoter. 
   
   
       23 . The method of  claim 17  wherein the protein of interest is a heterologous protein. 
   
   
       24 . The method of  claim 23  wherein the heterologous protein is selected from the group consisting of hormones, enzymes, growth factors, cytokines, and antibodies. 
   
   
       25 . The method of  claim 15  wherein the host cell is a filamentous fungus. 
   
   
       26 . The method of  claim 25  wherein the fungus is selected from the group consisting of  Trichoderma, Humicola, Fusarium, Aspergillus, Neurospora, Penicillium, Cephalosporium, Achlya, Podospora, Endothia, Mucor, Cochliobolus  and  Pyricularia.    
   
   
       27 . The method of  claim 26  wherein the fungus is  Trichoderma  spp. 
   
   
       28 . The method of  claim 27  wherein the fungus is  Trichoderma reesei.    
   
   
       29 . The method of  claim 26  wherein the fungus is  Penicillium  spp. 
   
   
       30 . The method of  claim 29  wherein the fungus is  Penicillium funiculosum.    
   
   
       31 . The method of  claim 15  wherein the host cell is a bacteria. 
   
   
       32 . The method of  claim 31  wherein the bacteria is selected from the group consisting of  Streptomyces, Thermomonospora, Bacillus,  and  Cellulomonas.    
   
   
       33 . The method of  claim 1  wherein the whole cellulase preparation is immobilized